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Sample records for nerve growth factor-induced

  1. Nerve growth factor induces sensitization of nociceptors without evidence for increased intraepidermal nerve fiber density.

    PubMed

    Hirth, Michael; Rukwied, Roman; Gromann, Alois; Turnquist, Brian; Weinkauf, Benjamin; Francke, Klaus; Albrecht, Philip; Rice, Frank; Hägglöf, Björn; Ringkamp, Matthias; Engelhardt, Maren; Schultz, Christian; Schmelz, Martin; Obreja, Otilia

    2013-11-01

    Nerve growth factor (NGF) is involved in the long-term sensitization of nociceptive processing linked to chronic pain. Functional and structural ("sprouting") changes can contribute. Thus, humans report long-lasting hyperalgesia to mechanical and electrical stimulation after intradermal NGF injection and NGF-induced sprouting has been reported to underlie cancer bone pain and visceral pain. Using a human-like animal model we investigated the relationship between the structure and function of unmyelinated porcine nociceptors 3 weeks after intradermal NGF treatment. Axonal and sensory characteristics were studied by in vivo single-fiber electrophysiology and immunohistochemistry. C fibers recorded extracellularly were classified based on mechanical response and activity-dependent slowing (ADS) of conduction velocity. Intraepidermal nerve fiber (IENF) densities were assessed by immunohistochemistry in pigs and in human volunteers using the same NGF model. NGF increased conduction velocity and reduced ADS and propagation failure in mechano-insensitive nociceptors. The proportion of mechano-sensitive C nociceptors within NGF-treated skin areas increased from 45.1% (control) to 71% and their median mechanical thresholds decreased from 40 to 20 mN. After NGF application, the mechanical receptive fields of nociceptors increased from 25 to 43 mm(2). At the structural level, however, IENF density was not increased by NGF. In conclusion, intradermal NGF induces long-lasting axonal and mechanical sensitization in porcine C nociceptors that corresponds to hyperalgesia observed in humans. Sensitization is not accompanied by increased IENF density, suggesting that NGF-induced hyperalgesia might not depend on changes in nerve fiber density but could be linked to the recruitment of previously silent nociceptors. Copyright © 2013 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

  2. Nerve growth factor induces the expression of chaperone protein calreticulin in human epithelial ovarian cells.

    PubMed

    Vera, C; Tapia, V; Kohan, K; Gabler, F; Ferreira, A; Selman, A; Vega, M; Romero, C

    2012-07-01

    Epithelial ovarian cancer is highly angiogenic and high expression of Nerve Growth Factor (NGF), a proangiogenic protein. Calreticulin is a multifunctional protein with anti-angiogenic properties and its translocation to the tumor cell membrane promotes recognition and engulfment by dendritic cells. The aim of this work was to evaluate calreticulin expression in human normal ovaries, benign and borderline tumors, and epithelial ovarian cancer samples and to evaluate whether NGF regulates calreticulin expression in human ovarian surface epithelium and in epithelial ovarian cancer cell lines. Calreticulin mRNA and protein levels were analyzed using RT-PCR, Western blot and immunohistochemistry in 67 human ovarian samples obtained from our Institution. Calreticulin expression induced by NGF stimulation in cell lines was evaluated using RT-PCR, Western blot and immunocytochemistry. We found a significant increase of calreticulin mRNA levels in epithelial ovarian cancer samples as compared to normal ovaries, benign tumors, and borderline tumors. Calreticulin protein levels, evaluated by Western blot, were also increased in epithelial ovarian cancer with respect to benign and borderline tumors. When HOSE and A2780 cell lines were stimulated with Nerve Growth Factor, we found an increase in calreticulin protein levels compared to controls. This effect was reverted by GW441756, a TRKA specific inhibitor. These results suggest that NGF regulates calreticulin protein levels in epithelial ovarian cells through TRKA receptor activation. © Georg Thieme Verlag KG Stuttgart · New York.

  3. Atlastin regulates store-operated calcium entry for nerve growth factor-induced neurite outgrowth

    PubMed Central

    Li, Jing; Yan, Bing; Si, Hongjiang; Peng, Xu; Zhang, Shenyuan L.; Hu, Junjie

    2017-01-01

    Homotypic membrane fusion of the endoplasmic reticulum (ER) is mediated by a class of dynamin-like GTPases known as atlastin (ATL). Depletion of or mutations in ATL cause an unbranched ER morphology and hereditary spastic paraplegia (HSP), a neurodegenerative disease characterized by axon shortening in corticospinal motor neurons and progressive spasticity of the lower limbs. How ER shaping is linked to neuronal defects is poorly understood. Here, we show that dominant-negative mutants of ATL1 in PC-12 cells inhibit nerve growth factor (NGF)-induced neurite outgrowth. Overexpression of wild-type or mutant ATL1 or depletion of ATLs alters ER morphology and affects store-operated calcium entry (SOCE) by decreasing STIM1 puncta formation near the plasma membrane upon calcium depletion of the ER. In addition, blockage of the STIM1-Orai pathway effectively abolishes neurite outgrowth of PC-12 cells stimulated by NGF. These results suggest that SOCE plays an important role in neuronal regeneration, and mutations in ATL1 may cause HSP, partly by undermining SOCE. PMID:28240257

  4. Nerve growth factor induced hyperalgesia in the rat hind paw is dependent on circulating neutrophils.

    PubMed

    Bennett, G; al-Rashed, S; Hoult, J R; Brain, S D

    1998-09-01

    The mechanisms by which nerve growth factor (NGF) induces thermal hyperalgesia and neutrophil accumulation have been investigated in the rat. Thermal nociceptive thresholds in rat hind paw were measured as the time taken for paw withdrawal from a heat source and neutrophil accumulation was measured in hind paw and dorsal skin samples using a myeloperoxidase assay. NGF (23-80 pmol intraplantar (i.pl.) injection) induced a significant (P < 0.05, n = 6-16) thermal hyperalgesia at 5 h after injection and significant neutrophil accumulation (P < 0.05, n = 6) was observed with NGF (40 pmol). In dorsal skin, where multiple samples can be assessed, intradermal (i.d.) NGF was 10-30 times less potent than interleukin-1beta in inducing neutrophil accumulation. The 5-lipoxygenase inhibitor ZM230487 (10 nmol co-injected with NGF) significantly attenuated neutrophil accumulation and hyperalgesia induced by NGF; unlike the histamine and 5-hydroxytryptamine antagonists (mepyramine and methysergide) which were without effect at the times measured. Furthermore, depletion of circulating neutrophils (using a rabbit anti-rat neutrophil antibody) abolished NGF induced hyperalgesia. These results indicate that neutrophils, which accumulate in response to a 5-lipoxygenase product, play a crucial role in NGF-induced hyperalgesia.

  5. Effect of a 5-lipoxygenase inhibitor on nerve growth factor-induced thermal hyperalgesia in the rat.

    PubMed

    Amann, R; Schuligoi, R; Lanz, I; Peskar, B A

    1996-06-13

    Intraplantar injection of mouse beta (2.5S) nerve growth factor (NGF) caused thermal hyperalgesia and stimulated release of immunoreactive leukotriene B4 from the rat paw skin. Both effects of NGF were prevented by the 5-lipoxygenase inhibitor, (R)-2-[4-quinolin-2-yl-methoxy)phenyl]-2-cyclopentyl acetic acid (BAY X1005). BAY X1005 did not affect bradykinin-induced thermal hyperalgesia. These results suggest the participation of 5-lipoxygenase products of arachidonate in NGF-induced local thermal hyperalgesia.

  6. Nerve growth factor-induced changes in neural cell adhesion molecule (N-CAM) in PC12 cells.

    PubMed Central

    Prentice, H M; Moore, S E; Dickson, J G; Doherty, P; Walsh, F S

    1987-01-01

    The effects of nerve growth factor (NGF) on the expression of neural cell adhesion molecule (N-CAM) in PC12 cells were determined. A quantitative immunoassay was used to show that NGF induces a 4- to 5-fold increase in relative N-CAM levels over a 3-day period. This increase could not be mimicked by cholera toxin suggesting that it is not a simple consequence of morphological differentiation. The changes in N-CAM levels induced by NGF were accompanied by changes in N-CAM molecular forms. The 140-kd N-CAM species is the major N-CAM expressed by naive PC12 cells, while NGF-treated cultures express N-CAM species of 180 kd and 140 kd. Northern analysis showed that naive cells express a 6.7-kd N-CAM mRNA species only, while NGF-treated cultures express both a 6.7-kb and a 7.2-kb transcript. As the 6.7-kb and 7.2-kb mRNAs are alternative spliced transcripts of a single gene, this result shows that NGF can activate a neuron-specific splicing mechanism. This is the first description of control of N-CAM expression by a growth factor. Images Fig. 3. Fig. 4. Fig. 5. PMID:3308447

  7. Lithium ion inhibits nerve growth factor-induced neurite outgrowth and phosphorylation of nerve growth factor-modulated microtubule-associated proteins

    PubMed Central

    1985-01-01

    LiCl (2.5-20 mM) reversibly suppressed nerve growth factor (NGF)- induced neurite outgrowth by cultured rat PC 12 pheochromocytoma cells. Similar concentrations of LiCl also reversibly blocked NGF-dependent regeneration of neurites by PC12 cells that had been primed by long- term pre-exposure to NGF and by cultured newborn mouse sympathetic neurons. In contrast, transcription-dependent responses of PC12 cells to NGF such as priming and induction of the NGF-inducible large external glycoprotein, occurred despite the presence of Li+. SDS PAGE analysis of total cellular phosphoproteins (labeled by 2-h exposure to 32P-orthophosphate) from neurite-bearing primed PC12 cells revealed that Li+ reversibly inhibited the phosphorylation of a band of Mr 64,000 that was barely detectable in NGF-untreated PC12 cells. However, Li+ did not appear to affect the labeling of other phosphoproteins in either NGF-primed or untreated PC12 cultures, nor did it affect the rapid increase in phosphorylation of several proteins that occurs when NGF is first added to unprimed cultures. Several criteria indicated that the NGF-inducible phosphoprotein of Mr 64,000 is a microtubule- associated protein (MAP). Of the NGF-inducible phosphorylated MAPs that have been detected in PC12 cells (Mr 64,000, 72,000, 80,000, and 320,000), several (Mr 64,000, 72,000, and 80,000) were found to be substantially less phosphorylated in the presence of Li+. Neither a phorbol ester tumor promotor nor permeant cAMP analogs reversed the inhibitory effects of Li+ on neurite outgrowth or on phosphorylation of the component of Mr 64,000. Microtubules are a major and required constituent of neurites, and MAPs may regulate the assembly and stability of neuritic microtubules. The observation that Li+ selectively inhibits NGF-induced neurite outgrowth and MAP phosphorylation suggests a possible causal relationship between these two events. PMID:4030895

  8. Intracerebroventricular administration of nerve growth factor induces gliogenesis in sensory ganglia, dorsal root, and within the dorsal root entry zone.

    PubMed

    Schlachetzki, Johannes C M; Pizzo, Donald P; Morrissette, Debbi A; Winkler, Jürgen

    2014-01-01

    Previous studies indicated that intracerebroventricular administration of nerve growth factor (NGF) leads to massive Schwann cell hyperplasia surrounding the medulla oblongata and spinal cord. This study was designed to characterize the proliferation of peripheral glial cells, that is, Schwann and satellite cells, in the trigeminal ganglia and dorsal root ganglia (DRG) of adult rats during two weeks of NGF infusion using bromodeoxyuridine (BrdU) to label dividing cells. The trigeminal ganglia as well as the cervical and lumbar DRG were analyzed. Along the entire neuraxis a small number of dividing cells were observed within these regions under physiological condition. NGF infusion has dramatically increased the generation of new cells in the neuronal soma and axonal compartments of sensory ganglia and along the dorsal root and the dorsal root entry zone. Quantification of BrdU positive cells within sensory ganglia revealed a 2.3- to 3-fold increase in glial cells compared to controls with a similar response to NGF for the different peripheral ganglia examined. Immunofluorescent labeling with S100β revealed that Schwann and satellite cells underwent mitosis after NGF administration. These data indicate that intracerebroventricular NGF infusion significantly induces gliogenesis in trigeminal ganglia and the spinal sensory ganglia and along the dorsal root entry zone as well as the dorsal root.

  9. Nerve growth factor-induced circadian phase shifts and MAP kinase activation in the hamster suprachiasmatic nuclei.

    PubMed

    Pizzio, Gastón A; Hainich, Ernesto C; Plano, Santiago A; Ralph, Martin R; Golombek, Diego A

    2005-08-01

    Circadian rhythms are entrained by light and by several neurochemical stimuli. In hamsters housed in constant darkness, i.c.v. administration of nerve growth factor (NGF) at various times in their circadian cycle produced phase shifts of locomotor activity rhythms that were similar in direction and circadian timing to those produced by brief pulses of light. Moreover, the effect of NGF and light were not additive, indicating signalling points in common. These points include the immediate-early gene c-fos and ERK1/2, a component of the mitogen-activated protein kinases (MAPK) family. NGF activates c-FOS and ERK1/2-MAPK in the suprachiasmatic nuclei, the site of a circadian clock in mammals, when administered during the subjective night but not during the day. The effect of NGF on ERK1/2 activation was not inhibited by the administration of MK-801, a glutamate/NMDA receptor antagonist. These results suggest that NGF, acting through MAPK activation, plays a role in photic entrainment of the mammalian circadian clock.

  10. Nerve growth factor induces rapid increases in functional cell surface low density lipoprotein receptor-related protein.

    PubMed

    Bu, G; Sun, Y; Schwartz, A L; Holtzman, D M

    1998-05-22

    The low density lipoprotein receptor-related protein (LRP) is a large endocytic receptor that binds multiple ligands and is highly expressed in neurons. Several LRP ligands, including apolipoprotein E/lipoproteins and amyloid precursor protein, have been shown to participate either in Alzheimer's disease pathogenesis or pathology. However, factors that regulate LRP expression in neurons are unknown. In the current study, we analyzed the effects of nerve growth factor (NGF) treatment on LRP expression, distribution, and function within neurons in two neuronal cell lines. Our results show that NGF induces a rapid increase of cell surface LRP expression in a central nervous system-derived neuronal cell line, GT1-1 Trk, which was seen within 10 min and reached a maximum at about 1 h of NGF treatment. This increase of cell surface LRP expression is concomitant with an increase in the endocytic activity of LRP as measured via ligand uptake and degradation assays. We also found that the cytoplasmic tail of LRP is phosphorylated and that NGF rapidly increases the amount of phosphorylation. Furthermore, we detected a significant increase of LRP expression at the messenger RNA level following 24 h of NGF treatment. Both rapid and long term induction of LRP expression were also detected in peripheral nervous system-derived PC12 cells following NGF treatment. Taken together, our results demonstrate that NGF regulates LRP expression in neuronal cells.

  11. Nerve growth factor-induced neurite sprouting in PC12 cells involves sigma-1 receptors: implications for antidepressants.

    PubMed

    Takebayashi, Minoru; Hayashi, Teruo; Su, Tsung-Ping

    2002-12-01

    One theory concerning the action of antidepressants relates to the drugs' ability to induce an adaptive plasticity in neurons such as neurite sprouting. Certain antidepressants are known to bind to sigma-1 receptors (Sig-1R) with high affinity. Sig-1R are dynamic endoplasmic reticulum proteins that are highly concentrated at the tip of growth cones in cultured cells. We therefore tested the hypotheses that Sig-1R might participate in the neurite sprouting and that antidepressants with Sig-1R affinity may promote the neuronal sprouting via Sig-1R. The prototypic Sig-1R agonist (+)-pentazocine [(+)PTZ], as well as the Sig-1R-active antidepressants imipramine and fluvoxamine, although ineffective by themselves, were found to enhance the nerve growth factor (NGF)-induced neurite sprouting in PC12 cells in a dose-dependent manner. A Sig-1R antagonist N,N-dipropyl-2-[4-methoxy-3-(2-phenylethoxy)phenyl]-ethylamine monohydrochloride (NE100) blocked the enhancements caused by these Sig-1R agonists. In separate experiments, we found that NGF dose and time dependently increased Sig-1R in PC12 cells. Chronic treatment of cells with (+)PTZ, imipramine, or fluvoxamine also increased Sig-1R. These latter results suggested that NGF induces the neurite sprouting by increasing Sig-1R. Indeed, the overexpression of Sig-1R per se in PC12 cells enhanced the NGF-induced neurite sprouting. Furthermore, antisense deoxyoligonucleotides directed against Sig-1R attenuated the NGF-induced neurite sprouting. Thus, when taken together, our results indicate that Sig-1R play an important role in the NGF-induced neurite sprouting and that certain antidepressants may facilitate neuronal sprouting in the brain via Sig-1R.

  12. NICER elements: a family of nerve growth factor-inducible cAMP-extinguishable retrovirus-like elements.

    PubMed Central

    Cho, K O; Minsk, B; Wagner, J A

    1990-01-01

    We have shown previously that the transcription of the gene designated d5 is induced by nerve growth factor (NGF) in rat adrenal pheochromocytoma PC-12 cells and that this NGF induction is repressed by cAMP. In this paper we demonstrate that d5 is a member of a gene family that contains several hundred members, which is closely related to retroviruses and retrotransposons, as demonstrated by the following observations: (i) the original d5 cDNA hybridized to numerous restriction fragments in genomic DNA; (ii) d5 cDNA hybridized to genomic clones with various intensities, and genomic clones can be isolated with a frequency suggesting that this family includes several hundred members; and (iii) there were minor sequence variations in four independently isolated cDNA clones that were homologous to d5 cDNA. Primer extension studies show that initiation of the 5.7-kilobase d5 mRNA(s) occurs at a unique site relative to a synthetic primer. The 5' end of the cDNA sequence was homologous to Rasheed rat sarcoma virus; and a genomic clone contained several elements that are typical of a long terminal repeat (LTR), including a CCAAT box, a TATA box, a primer binding site, a poly(A) addition signal, and a poly(A) addition site. Furthermore, there is a LTR at the 3' end of at least one of the genes in this family, and there appeared to be a four-base duplication at the probable site of integration into host DNA. Since several members of this family retain responses to NGF and cAMP, we conclude that the regulatory elements present in the LTR have been conserved in many members of this family. We have named this family of genes the NICER elements because they are a family of NGF-inducible cAMP-extinguishable retrovirus-like elements. Images PMID:2160077

  13. The transcription factor nerve growth factor-inducible protein a mediates epigenetic programming: altering epigenetic marks by immediate-early genes.

    PubMed

    Weaver, Ian C G; D'Alessio, Ana C; Brown, Shelley E; Hellstrom, Ian C; Dymov, Sergiy; Sharma, Shakti; Szyf, Moshe; Meaney, Michael J

    2007-02-14

    Maternal care alters epigenetic programming of glucocorticoid receptor (GR) gene expression in the hippocampus, and increased postnatal maternal licking/grooming (LG) behavior enhances nerve growth factor-inducible protein A (NGFI-A) transcription factor binding to the exon 1(7) GR promoter within the hippocampus of the offspring. We tested the hypothesis that NGFI-A binding to the exon 1(7) GR promoter sequence marks this sequence for histone acetylation and DNA demethylation and that such epigenetic alterations subsequently influence NGFI-A binding and GR transcription. We report that (1) NGFI-A binding to its consensus sequence is inhibited by DNA methylation, (2) NGFI-A induces the activity of exon 1(7) GR promoter in a transient reporter assay, (3) DNA methylation inhibits exon 1(7) GR promoter activity, and (4) whereas NGFI-A interaction with the methylated exon 1(7) GR promoter is reduced, NGFI-A overexpression induces histone acetylation, DNA demethylation, and activation of the exon 1(7) GR promoter in transient transfection assays. Site-directed mutagenesis assays demonstrate that NGFI-A binding to the exon 1(7) GR promoter is required for such epigenetic reprogramming. In vivo, enhanced maternal LG is associated with increased NGFI-A binding to the exon 1(7) GR promoter in the hippocampus of pups, and NGFI-A-bound exon 1(7) GR promoter is unmethylated compared with unbound exon 1(7) GR promoter. Knockdown experiments of NGFI-A in hippocampal primary cell culture show that NGFI-A is required for serotonin-induced DNA demethylation and increased exon 1(7) GR promoter expression. The data are consistent with the hypothesis that NGFI-A participates in epigenetic programming of GR expression.

  14. The ras suppressor, RSU-1, enhances nerve growth factor-induced differentiation of PC12 cells and induces p21CIP expression.

    PubMed

    Masuelli, L; Ettenberg, S; Vasaturo, F; Vestergaard-Sykes, K; Cutler, M L

    1999-08-01

    The Rsu-1 Ras suppressor gene was isolated based on its ability to inhibit v-Ras transformation. Using Rsu-1 transfectants of the pheochromocytoma cell line PC12, we demonstrated previously that Rsu-1 expression inhibited Jun kinase activation but enhanced Erk2 activation in response to epidermal growth factor. In the present study, the Rsu-1 PC12 transfectants were used to investigate the role of Rsu-1 in nerve growth factor (NGF)- and v-Ki-ras-mediated neuronal differentiation. NGF-induced neurite extension was enhanced, not inhibited, by the expression of Rsu-1 in PC12 cells. The activation of Erk kinase activity in response to NGF was sustained longer in the Rsu-1 transfectants compared with the vector control cells. During NGF-mediated differentiation, an increase in the expression of specific mRNAs for the early response genes Fos, cJun, and NGF1a was detected in both the vector control and Rsu-1 transfectants. The expression of the differentiation-specific genes VGF8 and SCG10 was similar in Rsu-1 transfectants compared with the vector control cells. The induction of Rsu-1 expression in these cell lines did not inhibit v-Ki-ras-induced differentiation, as measured by neurite extension. These data suggest that although Rsu-1 blocked some Ras-dependent response(s), these responses were not required for differentiation. Moreover, the induction of Rsu-1 expression in the PC12 clones resulted in growth inhibition and p21(WAF/CIP) expression. Hence, Rsu-1 expression enhances NGF-induced differentiation while inhibiting the growth of cells.

  15. Nerve growth factor-induced changes in the intracellular localization of the protein kinase C substrate B-50 in pheochromocytoma PC12 cells

    PubMed Central

    1989-01-01

    High levels of the neuron-specific protein kinase C substrate, B-50 (= GAP43), are present in neurites and growth cones during neuronal development and regeneration. This suggests a hitherto nonelucidated role of this protein in neurite outgrowth. Comparable high levels of B- 50 arise in the pheochromocytoma PC12 cell line during neurite formation. To get insight in the putative growth-associated function of B-50, we compared its ultrastructural localization in naive PC12 cells with its distribution in nerve growth factor (NGF)- or dibutyryl cyclic AMP (dbcAMP)-treated PC12 cells. B-50 immunogold labeling of cryosections of untreated PC12 cells is mainly associated with lysosomal structures, including multivesicular bodies, secondary lysosomes, and Golgi apparatus. The plasma membrane is virtually devoid of label. However, after 48-h NGF treatment of the cells, B-50 immunoreactivity is most pronounced on the plasma membrane. Highest B- 50 immunoreactivity is observed on plasma membranes surrounding sprouting microvilli, lamellipodia, and filopodia. Outgrowing neurites are scattered with B-50 labeling, which is partially associated with chromaffin granules. In NGF-differentiated PC12 cells, B-50 immunoreactivity is, as in untreated cells, also associated with organelles of the lysosomal family and Golgi stacks. B-50 distribution in dbcAMP-differentiated cells closely resembles that in NGF-treated cells. The altered distribution of B-50 immunoreactivity induced by differentiating agents indicates a shift of the B-50 protein towards the plasma membrane. This translocation accompanies the acquisition of neuronal features of PC12 cells and points to a neurite growth- associated role for B-50, performed at the plasma membrane at the site of protrusion. PMID:2537833

  16. Distinctive effect on nerve growth factor-induced PC12 cell neurite outgrowth by two unique neolignan enantiomers from Illicium merrillianum

    NASA Astrophysics Data System (ADS)

    Tian, Xinhui; Yue, Rongcai; Zeng, Huawu; Li, Honglin; Shan, Lei; He, Weiwei; Shen, Yunheng; Zhang, Weidong

    2015-11-01

    Merrillianoid (1), a racemic neolignan possessing the characteristic benzo-2,7-dioxabicyclo[3.2.1]octane moiety, was isolated from the branches and leaves of Illicium merrillianum. Chiral separation of 1 gave two enantiomers (+)-1 and (-)-1. The structure of 1 was established by comprehensive spectroscopic analysis and single crystal X-ray diffraction. The absolute configurations of enantiomers were determined by quantum mechanical calculation. Compound (+)-1 exhibited a better neurotrophic activity than racemate 1 by promoting nerve growth factor (NGF) induced PC12 cell neurite outgrowth, while (-)-1 showed a distinctive inhibitory effect. Furthermore, a mechanism study indicated that the two enantiomers influenced NGF-induced neurite outgrowth of PC12 cells possibly by interacting with the trkA receptor, and extracellular signal regulated kinases 1/2 (ERK1/2) and mitogen-activated protein kinase (MEK) in Ras/ERK signal cascade. But the phosphorylation level of serine/threonine kinase Akt1 and Akt2 in PI3K/Akt signal pathway showed no significant difference between (+)-1 and (-)-1.

  17. Pro-nerve Growth Factor Induces Autocrine Stimulation of Breast Cancer Cell Invasion through Tropomyosin-related Kinase A (TrkA) and Sortilin Protein*

    PubMed Central

    Demont, Yohann; Corbet, Cyril; Page, Adeline; Ataman-Önal, Yasemin; Choquet-Kastylevsky, Genevieve; Fliniaux, Ingrid; Le Bourhis, Xuefen; Toillon, Robert-Alain; Bradshaw, Ralph A.; Hondermarck, Hubert

    2012-01-01

    The precursor of nerve growth factor (proNGF) has been described as a biologically active polypeptide able to induce apoptosis in neuronal cells, via the neurotrophin receptor p75NTR and the sortilin receptor. Herein, it is shown that proNGF is produced and secreted by breast cancer cells, stimulating their invasion. Using Western blotting and mass spectrometry, proNGF was detected in a panel of breast cancer cells as well as in their conditioned media. Immunohistochemical analysis indicated an overproduction of proNGF in breast tumors, when compared with benign and normal breast biopsies, and a relationship to lymph node invasion in ductal carcinomas. Interestingly, siRNA against proNGF induced a decrease of breast cancer cell invasion that was restored by the addition of non-cleavable proNGF. The activation of TrkA, Akt, and Src, but not the MAP kinases, was observed. In addition, the proNGF invasive effect was inhibited by the Trk pharmacological inhibitor K252a, a kinase-dead TrkA, and siRNA against TrkA sortilin, neurotensin, whereas siRNA against p75NTR and the MAP kinase inhibitor PD98059 had no impact. These data reveal the existence of an autocrine loop stimulated by proNGF and mediated by TrkA and sortilin, with the activation of Akt and Src, for the stimulation of breast cancer cell invasion. PMID:22128158

  18. Asarone from Acori Tatarinowii Rhizoma Potentiates the Nerve Growth Factor-Induced Neuronal Differentiation in Cultured PC12 Cells: A Signaling Mediated by Protein Kinase A

    PubMed Central

    Lam, Kelly Y. C.; Chen, Jianping; Lam, Candy T. W.; Wu, Qiyun; Yao, Ping; Dong, Tina T. X.; Lin, Huangquan; Tsim, Karl W. K.

    2016-01-01

    Acori Tatarinowii Rhizoma (ATR), the rhizome of Acorus tatarinowii Schott, is being used clinically to treat neurological disorders. The volatile oil of ATR is being considered as an active ingredient. Here, α-asarone and β-asarone, accounting about 95% of ATR oil, were evaluated for its function in stimulating neurogenesis. In cultured PC12 cells, application of ATR volatile oil, α-asarone or β-asarone, stimulated the expression of neurofilaments, a bio-marker for neurite outgrowth, in a concentration-dependent manner. The co-treatment of ATR volatile oil, α-asarone or β-asarone, with low concentration of nerve growth factor (NGF) potentiated the NGF-induced neuronal differentiation in cultured PC12 cells. In addition, application of protein kinase A inhibitors, H89 and KT5720, in cultures blocked the ATR-induced neurofilament expression, as well as the phosphorylation of cAMP-responsive element binding protein (CREB). In the potentiation of NGF-induced signaling in cultured PC12 cells, α-asarone and β-asarone showed synergistic effects. These results proposed the neurite-promoting asarone, or ATR volatile oil, could be useful in finding potential drugs for treating various neurodegenerative diseases, in which neurotrophin deficiency is normally involved. PMID:27685847

  19. ERK5 Activity Is Required for Nerve Growth Factor-induced Neurite Outgrowth and Stabilization of Tyrosine Hydroxylase in PC12 Cells*

    PubMed Central

    Obara, Yutaro; Yamauchi, Arata; Takehara, Shin; Nemoto, Wataru; Takahashi, Maho; Stork, Philip J. S.; Nakahata, Norimichi

    2009-01-01

    Extracellular signal-regulated kinases (ERKs) play important physiological roles in proliferation, differentiation, and gene expression. ERK5 is approximately twice the size of ERK1/2, and its amino-terminal half contains the kinase domain that shares homology with ERK1/2 and TEY activation motif, whereas the carboxyl-terminal half is unique. In this study, we examined a physiological role of ERK5 in rat pheochromocytoma cells (PC12), comparing it with ERK1/2. Nerve growth factor (NGF) induced phosphorylation of both ERK5 and ERK1/2, whereas the cAMP analog dibutyryl cAMP (Bt2cAMP) caused only ERK1/2 phosphorylation. U0126, at 30 μm, that blocks ERK1/2 signaling selectively attenuated neurite outgrowth induced by NGF and Bt2cAMP, but BIX02188 and BIX02189, at 30 μm, that block ERK5 signaling and an ERK5 dominant-negative mutant suppressed only NGF-induced neurite outgrowth. Next, we examined the expression of tyrosine hydroxylase, a rate-limiting enzyme of catecholamine biosynthesis. Both NGF and Bt2cAMP increased tyrosine hydroxylase gene promoter activity in an ERK1/2-dependent manner but was ERK5-independent. However, when both ERK5 and ERK1/2 signalings were inhibited, tyrosine hydroxylase protein up-regulation by NGF and Bt2cAMP was abolished, because of the loss of stabilization of tyrosine hydroxylase protein by ERK5. Taking these results together, ERK5 is involved in neurite outgrowth and stabilization of tyrosine hydroxylase in PC12 cells, and ERK5, along with ERK1/2, plays essential roles in the neural differentiation process. PMID:19581298

  20. Nerve growth factor induced changes in the Golgi apparatus of PC-12 rat pheochromocytoma cells as studied by ligand endocytosis, cytochemical and morphometric methods.

    PubMed

    Hickey, W F; Stieber, A; Hogue-Angeletti, R; Gonatas, J; GOnatas, N K

    1983-10-01

    Cells of the PC-12 rat pheochromocytoma cell line respond to nerve growth factor (NGF) by sprouting neurites and biochemically differentiating into sympathetic ganglion-like cells. NGF-stimulated ('differentiated') and unstimulated ('undifferentiated') cells were studied by cytochemical techniques for the localization of the enzymes acid phosphatase (ACPase) and thiamine pyrophosphatase (TPPase), and by a morphometric analysis of the distribution of endocytosed wheat-germ agglutinin labelled with horseradish peroxidase (WGA-HRP). Both cytochemical stains showed the enzymes to be distributed in lysosomes and certain cisternae of the Golgi apparatus in both NGF stimulated and unstimulated cells. ACPase was not confined to GERL (Golgi-endoplasmic reticulum-lysosome) as in certain other cells. The morphometric studies demonstrated that the reaction product of the internalized WGA-HRP occupied 4.7% of the cytoplasmic area in unstimulated cells and 4.5% in NGF-stimulated ones. Despite this similarity, the distribution of the WGA-HRP among the studied intracellular compartments in these two cell groups varied. In the NGF-stimulated cells 3.3% of the WGA-HRP reaction product was found in the innermost Golgi cisterna(e) while in unstimulated cells only 0.3% was seen in this compartment. Similarly, 4.3% of the WGA-HRP stain was found in small vesicles at the 'trans' aspect of the Golgi apparatus in stimulated cells, when only 0.3% of the stain occupied this compartment in 'undifferentiated' cells. The morphometric analysis also revealed that when the PC-12 cells were stimulated with NGF, the Golgi apparatus increased in area by approximately 70%. These findings are consistent with the hypothesis that NGF induced differentiation of PC-12 cells is coupled with enhanced endocytosis of WGA and probably of its 'receptor' to the innermost Golgi cisterna(e) and the closely associated vesicles.

  1. P2X1 Receptor-Mediated Ca(2+) Influx Triggered by DA-9801 Potentiates Nerve Growth Factor-Induced Neurite Outgrowth.

    PubMed

    Back, Moon Jung; Lee, Hae Kyung; Lee, Joo Hyun; Fu, Zhicheng; Son, Mi Won; Choi, Sang Zin; Go, Hyo Sang; Yoo, Sungjae; Hwang, Sun Wook; Kim, Dae Kyong

    2016-11-16

    Nerve growth factor (NGF)-induced neuronal regeneration has emerged as a strategy to treat neuronal degeneration-associated disorders. However, direct NGF administration is limited by the occurrence of adverse effects at high doses of NGF. Therefore, development of a therapeutic strategy to promote the NGF trophic effect is required. In view of the lack of understanding of the mechanism for potentiating the NGF effect, this study investigated molecular targets of DA-9801, a well-standardized Dioscorea rhizome extract, which has a promoting effect on NGF. An increase in intracellular calcium ion level was induced by DA-9801, and chelation of extracellular calcium ions with ethylene-bis(oxyethylenenitrilo)tetraacetic acid (EGTA) suppressed the potentiating effect of DA-9801 on NGF-induced neurite outgrowth. In addition, EGTA treatment reduced the DA-9801-induced phosphorylation of extracellular signal-regulated kinase1/2 (ERK1/2), the major mediators of neurite outgrowth. To find which calcium ion-permeable channel contributes to the calcium ion influx induced by DA-9801, we treated PC12 cells with various inhibitors of calcium ion-permeable channels. NF449, a P2X1 receptor selective antagonist, significantly abolished the potentiating effect of DA-9801 on NGF-induced neurite outgrowth and abrogated the DA-9801-induced ERK1/2 phosphorylation. In addition, transfection with siRNA of P2X1 receptor significantly reduced the DA-9801-enhanced neurite outgrowth. In conclusion, calcium ion influx through P2X1 receptor mediated the promoting effect of DA-9801 on NGF-induced neurite outgrowth via ERK1/2 phosphorylation.

  2. Potentiation of Nerve Growth Factor-Induced Neurite Outgrowth by Fluvoxamine: Role of Sigma-1 Receptors, IP3 Receptors and Cellular Signaling Pathways

    PubMed Central

    Nishimura, Tomoko; Ishima, Tamaki; Iyo, Masaomi; Hashimoto, Kenji

    2008-01-01

    Background Selective serotonin reuptake inhibitors (SSRIs) have been widely used and are a major therapeutic advance in psychopharmacology. However, their pharmacology is quite heterogeneous. The SSRI fluvoxamine, with sigma-1 receptor agonism, is shown to potentiate nerve-growth factor (NGF)-induced neurite outgrowth in PC 12 cells. However, the precise cellular and molecular mechanisms underlying potentiation by fluvoxamine are not fully understood. In this study, we examined the roles of cellular signaling pathways in the potentiation of NGF-induced neurite outgrowth by fluvoxamine and sigma-1 receptor agonists. Methods and Findings The effects of three SSRIs (fluvoxamine, sertraline, paroxetine) and three sigma-1 receptor agonists (SA4503, 4-phenyl-1-(4-phenylbutyl) piperidine (PPBP), and dehydroepiandrosterone (DHEA)-sulfate) on NGF-induced neurite outgrowth in PC12 cells were examined. Also examined were the effects of the sigma-1 receptor antagonist NE-100, inositol 1,4,5-triphosphate (IP3) receptor antagonist, and specific inhibitors of signaling pathways in the potentiation of NGF-induced neurite outgrowth by selective sigma-1 receptor agonist SA4503. Fluvoxamine (but not sertraline or paroxetine) and the sigma-1 receptor agonists SA4503, PPBP, and DHEA-sulfate significantly potentiated NGF-induced neurite outgrowth in PC12 cells in a concentration-dependent manner. The potentiation by fluvoxamine and the three sigma-1 receptor agonists was blocked by co-administration of the selective sigma-1 receptor antagonist NE-100, suggesting that sigma-1 receptors play a role in blocking the enhancement of NGF-induced neurite outgrowth. Moreover, the potentiation by SA4503 was blocked by co-administration of the IP3 receptor antagonist xestospongin C. In addition, the specific inhibitors of phospholipase C (PLC-γ), phosphatidylinositol 3-kinase (PI3K), p38MAPK, c-Jun N-terminal kinase (JNK), and the Ras/Raf/mitogen-activated protein kinase (MAPK) signaling pathways

  3. Postnatal expression of the plasticity-related nerve growth factor-induced gene A (NGFI-A) protein in the superficial layers of the rat superior colliculus: relation to N-methyl-D-aspartate receptor function.

    PubMed

    Giraldi-Guimarães, A; de Bittencourt-Navarrete, R E; Nascimento, I C C; Salazar, P R; Freitas-Campos, D; Mendez-Otero, R

    2004-01-01

    Immediate early gene expression in the CNS is induced by sensory stimulation and seems to be involved in long-term synaptic plasticity. We have used an immunohistochemical method to detect the nerve growth factor-induced gene A (NGFI-A) protein expression in the superficial layers of the rat superior colliculus during postnatal development. Our goal was to correlate the expression of this candidate plasticity protein with developmental events, especially the activity-dependent refinement of the retinocollicular and corticocollicular pathways. We have also investigated the N-methyl-D-aspartate (NMDA)-receptor dependence of the NGFI-A expression. Animals of various postnatal ages were used. Postnatal day (P) 12 and older animals were submitted to a protocol of dark adaptation followed by light stimulation. NGFI-A expression was never observed during the first 2 postnatal weeks. The first stained cells were observed at P15, 2 days after eye opening (P13). The highest number of stained cells was observed at the end of the third postnatal week (P22). Adult-like level of expression was reached at P30, since at this age, the number of stained cells was comparable to that found in adult rats (P90). Both P22 animals submitted to an acute treatment with MK-801 (i.p. injection) and adult animals submitted to chronic intracranial infusion of a MK-801 presented a clear decrease in the NGFI-A expression in response to light stimulation. These results suggest that the NGFI-A expression is dependent on the NMDA receptor activation, and the observed pattern of expression is in close agreement with previous descriptions of the changes in the NMDA receptor-mediated visual activity in the developing rat superior colliculus (SC). Our results suggest that the plasticity-related NGFI-A protein might play a role in the developmental plasticity of the superficial layers of the rat SC after eye opening.

  4. Cbl competitively inhibits epidermal growth factor-induced activation of phospholipase C-gamma1.

    PubMed

    Choi, Jang Hyun; Bae, Sun Sik; Park, Jong Bae; Ha, Sang Hoon; Song, Hebok; Kim, Jae-Ho; Cocco, Lucio; Ryu, Sung Ho; Suh, Pann-Ghill

    2003-04-30

    Phospholipase C-gamma1 (PLC-gamma1) plays pivotal roles in cellular growth and proliferation through its two Src homology (SH) 2 domains and its single SH3 domain, which interact with signaling molecules in response to various growth factors and hormones. However, the role of the SH domains in the growth factor-induced regulation of PLC-gamma1 is unclear. By peptide-mass fingerprinting analysis we have identified Cbl as a binding protein for the SH3 domain of PLC-gamma1 from rat pheochromatocyte PC12 cells. Association of Cbl with PLC-gamma1 was induced by epidermal growth factor (EGF) but not by nerve growth factor (NGF). Upon EGF stimulation, both Cbl and PLC-gamma1 were recruited to the activated EGF receptor through their SH2 domains. Mutation of the SH2 domains of either Cbl or PLC-gamma1 abrogated the EGF-induced interaction of PLC-gamma1 with Cbl, indicating that SH2-mediated translocation is essential for the association of PLC-gamma1 and Cbl. Overexpression of Cbl attenuated EGF-induced tyrosine phosphorylation and the subsequent activation of PLC-gamma1 by interfering competitively with the interaction between PLC-gamma1 and EGFR. Taken together, these results provide the first indications that Cbl may be a negative regulator of intracellular signaling following EGF-induced PLC-gamma1 activation.

  5. Basic Fibroblast Growth Factor Induces Angiogenesis in vitro

    NASA Astrophysics Data System (ADS)

    Montesano, R.; Vassalli, J.-D.; Baird, A.; Guillemin, R.; Orci, L.

    1986-10-01

    Fibroblast growth factors (FGFs) are potent mitogens for vascular and capillary endothelial cells in vitro and can stimulate the formation of blood capillaries (angiogenesis) in vivo. A crucial event in this process is the invasion of the perivascular extracellular matrix by sprouting endothelial cells. Using a recently developed in vitro model of angiogenesis, we show here that highly purified basic pituitary FGF can induce capillary endothelial cells to invade a three-dimensional collagen matrix and to organize themselves to form characteristic tubules that resemble blood capillaries. We also show that basic FGF concomitantly stimulates endothelial cells to produce a urokinase-type plasminogen activator, a protease that has been implicated in the neovascular response. The results demonstrate that basic FGF can stimulate processes that are characteristic of angiogenesis in vivo, including endothelial cell migration, invasion, and production of plasminogen activator.

  6. Keratinocyte growth factor induces pancreatic ductal epithelial proliferation.

    PubMed

    Yi, E S; Yin, S; Harclerode, D L; Bedoya, A; Bikhazi, N B; Housley, R M; Aukerman, S L; Morris, C F; Pierce, G F; Ulich, T R

    1994-07-01

    Keratinocyte growth factor (KGF) causes a proliferation of pancreatic ductal epithelial cells in adult rats after daily systemic administration for 1 to 2 weeks. Even before the proliferation of intralobular ducts is histologically evident, KGF also induces proliferating cell nuclear antigen expression within the ductal epithelium of intercalated, intralobular, and interlobular ducts. KGF also causes incorporation of 5-bromodeoxyuridine in ductal epithelial cells. Epithelial cell proliferation is histologically most prominent at the level of the intralobular ducts adjacent to and within the islets of Langerhans. Pancreatic ductal proliferation is not histologically apparent in rats sacrificed 7 to 10 days after the cessation of KGF administration. The pancreatic hormones insulin, glucagon, somatostatin, and pancreatic polypeptide are normally distributed within islets that demonstrate intrainsular ductal proliferation. The proliferating ductal epithelium does not show endocrine differentiation as evidenced by the lack of immunoreactivity for pancreatic hormones. KGF is a potent in vivo mitogen for pancreatic ductal epithelial cells.

  7. Connective tissue growth factor induces cardiac hypertrophy through Akt signaling

    SciTech Connect

    Hayata, Nozomi; Fujio, Yasushi; Yamamoto, Yasuhiro; Iwakura, Tomohiko; Obana, Masanori; Takai, Mika; Mohri, Tomomi; Nonen, Shinpei; Maeda, Makiko; Azuma, Junichi

    2008-05-30

    In the process of cardiac remodeling, connective tissue growth factor (CTGF/CCN2) is secreted from cardiac myocytes. Though CTGF is well known to promote fibroblast proliferation, its pathophysiological effects in cardiac myocytes remain to be elucidated. In this study, we examined the biological effects of CTGF in rat neonatal cardiomyocytes. Cardiac myocytes stimulated with full length CTGF and its C-terminal region peptide showed the increase in cell surface area. Similar to hypertrophic ligands for G-protein coupled receptors, such as endothelin-1, CTGF activated amino acid uptake; however, CTGF-induced hypertrophy is not associated with the increased expression of skeletal actin or BNP, analyzed by Northern-blotting. CTGF treatment activated ERK1/2, p38 MAPK, JNK and Akt. The inhibition of Akt by transducing dominant-negative Akt abrogated CTGF-mediated increase in cell size, while the inhibition of MAP kinases did not affect the cardiac hypertrophy. These findings indicate that CTGF is a novel hypertrophic factor in cardiac myocytes.

  8. Epidermal Growth Factor-induced Vacuolar (H+)-ATPase Assembly

    PubMed Central

    Xu, Yanqing; Parmar, Amanda; Roux, Emmanuelle; Balbis, Alejandro; Dumas, Victor; Chevalier, Stephanie; Posner, Barry I.

    2012-01-01

    Using proteomics and immunofluorescence, we demonstrated epidermal growth factor (EGF) induced recruitment of extrinsic V1 subunits of the vacuolar (H+)-ATPase (V-ATPase) to rat liver endosomes. This was accompanied by reduced vacuolar pH. Bafilomycin, an inhibitor of V-ATPase, inhibited EGF-stimulated DNA synthesis and mammalian target of rapamycin complex 1 (mTORC1) activation as indicated by a decrease in eukaryotic initiation factor 4E-binding 1 (4E-BP1) phosphorylation and p70 ribosomal S6 protein kinase (p70S6K) phosphorylation and kinase activity. There was no corresponding inhibition of EGF-induced Akt and extracellular signal-regulated kinase (Erk) activation. Chloroquine, a neutralizer of vacuolar pH, mimicked bafilomycin effects. Bafilomycin did not inhibit the association of mTORC1 with Raptor nor did it affect AMP-activated protein kinase activity. Rather, the intracellular concentrations of essential but not non-essential amino acids were decreased by bafilomycin in EGF-treated primary rat hepatocytes. Cycloheximide, a translation elongation inhibitor known to augment intracellular amino acid levels, prevented the effect of bafilomycin on amino acids levels and completely reversed its inhibition of EGF-induced mTORC1 activation. In vivo administration of EGF stimulated the recruitment of Ras homologue enriched in brain (Rheb) but not mammalian target of rapamycin (mTOR) to endosomes and lysosomes. This was inhibited by chloroquine treatment. Our results suggest a role for vacuolar acidification in EGF signaling to mTORC1. PMID:22689575

  9. Obesity and Postmenopausal Breast Cancer Risk: Determining the Role of Growth Factor-Induced Aromatase Expression

    DTIC Science & Technology

    2013-01-01

    Moreau T, Senouci K, Salmon RJ, Fourquet A, Asselain B. Is obesity an independent prognosis factor in woman breast cancer ? Breast Cancer Res Treat...1 AD__________________________ Award number: W81XWH-11-1-0132 Title: Obesity and Postmenopausal Breast Cancer Risk: Determining the Role... Obesity and Postmenopausal Breast Cancer Risk: Determining the Role of Growth Factor-Induced Aromatase Expression 5a. CONTRACT NUMBER 5b. GRANT

  10. Bone morphogenetic protein-4 strongly potentiates growth factor-induced proliferation of mammary epithelial cells

    SciTech Connect

    Montesano, Roberto Sarkoezi, Rita; Schramek, Herbert

    2008-09-12

    Bone morphogenetic proteins (BMPs) are multifunctional cytokines that elicit pleiotropic effects on biological processes such as cell proliferation, cell differentiation and tissue morphogenesis. With respect to cell proliferation, BMPs can exert either mitogenic or anti-mitogenic activities, depending on the target cells and their context. Here, we report that in low-density cultures of immortalized mammary epithelial cells, BMP-4 did not stimulate cell proliferation by itself. However, when added in combination with suboptimal concentrations of fibroblast growth factor (FGF)-2, FGF-7, FGF-10, epidermal growth factor (EGF) or hepatocyte growth factor (HGF), BMP-4 potently enhanced growth factor-induced cell proliferation. These results reveal a hitherto unsuspected interplay between BMP-4 and growth factors in the regulation of mammary epithelial cell proliferation. We suggest that the ability of BMP-4 to potentiate the mitogenic activity of multiple growth factors may contribute to mammary gland ductal morphogenesis as well as to breast cancer progression.

  11. Exposure to Nerve Growth Factor Worsens Nephrotoxic Effect Induced by Cyclosporine A in HK-2 Cells

    PubMed Central

    Lofaro, Danilo; Toteda, Giuseppina; Lupinacci, Simona; Leone, Francesca; Gigliotti, Paolo; Papalia, Teresa; Bonofiglio, Renzo

    2013-01-01

    Nerve growth factor is a neurotrophin that promotes cell growth, differentiation, survival and death through two different receptors: TrkANTR and p75NTR. Nerve growth factor serum concentrations increase during many inflammatory and autoimmune diseases, glomerulonephritis, chronic kidney disease, end-stage renal disease and, particularly, in renal transplant. Considering that nerve growth factor exerts beneficial effects in the treatment of major central and peripheral neurodegenerative diseases, skin and corneal ulcers, we asked whether nerve growth factor could also exert a role in Cyclosporine A-induced graft nephrotoxicity. Our hypothesis was raised from basic evidence indicating that Cyclosporine A-inhibition of calcineurin-NFAT pathway increases nerve growth factor expression levels. Therefore, we investigated the involvement of nerve growth factor and its receptors in the damage exerted by Cyclosporine A in tubular renal cells, HK-2. Our results showed that in HK-2 cells combined treatment with Cyclosporine A + nerve growth factor induced a significant reduction in cell vitality concomitant with a down-regulation of Cyclin D1 and up-regulation of p21 levels respect to cells treated with Cyclosporine A alone. Moreover functional experiments showed that the co-treatment significantly up-regulated human p21promoter activity by involvement of the Sp1 transcription factor, whose nuclear content was negatively regulated by activated NFATc1. In addition we observed that the combined exposure to Cyclosporine A + nerve growth factor promoted an up-regulation of p75 NTR and its target genes, p53 and BAD leading to the activation of intrinsic apoptosis. Finally, the chemical inhibition of p75NTR down-regulated the intrinsic apoptotic signal. We describe two new mechanisms by which nerve growth factor promotes growth arrest and apoptosis in tubular renal cells exposed to Cyclosporine A. PMID:24244623

  12. Acetylation of RNA polymerase II regulates growth-factor-induced gene transcription in mammalian cells.

    PubMed

    Schröder, Sebastian; Herker, Eva; Itzen, Friederike; He, Daniel; Thomas, Sean; Gilchrist, Daniel A; Kaehlcke, Katrin; Cho, Sungyoo; Pollard, Katherine S; Capra, John A; Schnölzer, Martina; Cole, Philip A; Geyer, Matthias; Bruneau, Benoit G; Adelman, Karen; Ott, Melanie

    2013-11-07

    Lysine acetylation regulates transcription by targeting histones and nonhistone proteins. Here we report that the central regulator of transcription, RNA polymerase II, is subject to acetylation in mammalian cells. Acetylation occurs at eight lysines within the C-terminal domain (CTD) of the largest polymerase subunit and is mediated by p300/KAT3B. CTD acetylation is specifically enriched downstream of the transcription start sites of polymerase-occupied genes genome-wide, indicating a role in early stages of transcription initiation or elongation. Mutation of lysines or p300 inhibitor treatment causes the loss of epidermal growth-factor-induced expression of c-Fos and Egr2, immediate-early genes with promoter-proximally paused polymerases, but does not affect expression or polymerase occupancy at housekeeping genes. Our studies identify acetylation as a new modification of the mammalian RNA polymerase II required for the induction of growth factor response genes.

  13. Nerve growth factor released from a novel PLGA nerve conduit can improve axon growth

    NASA Astrophysics Data System (ADS)

    Lin, Keng-Min; Shea, Jill; Gale, Bruce K.; Sant, Himanshu; Larrabee, Patti; Agarwal, Jay

    2016-04-01

    Nerve injury can occur due to penetrating wounds, compression, traumatic stretch, and cold exposure. Despite prompt repair, outcomes are dismal. In an attempt to help resolve this challenge, in this work, a poly-lactic-co-glycolic acid (PLGA) nerve conduit with associated biodegradable drug reservoir was designed, fabricated, and tested. Unlike current nerve conduits, this device is capable of fitting various clinical scenarios by delivering different drugs without reengineering the whole system. To demonstrate the potential of this device for nerve repair, a series of experiments were performed using nerve growth factor (NGF). First, an NGF dosage curve was developed to determine the minimum NGF concentration for optimal axonal outgrowth on chick dorsal root ganglia (DRG) cells. Next, PLGA devices loaded with NGF were evaluated for sustained drug release and axon growth enhancement with the released drug. A 20 d in vitro release test was conducted and the nerve conduit showed the ability to meet and maintain the minimum NGF requirement determined previously. Bioactivity assays of the released NGF showed that drug released from the device between the 15th and 20th day could still promote axon growth (76.6-95.7 μm) in chick DRG cells, which is in the range of maximum growth. These novel drug delivery conduits show the ability to deliver NGF at a dosage that efficiently promotes ex vivo axon growth and have the potential for in vivo application to help bridge peripheral nerve gaps.

  14. [Progress of study on inhibitory effects of traditional Chinese herbs on growth factor induced proliferation of vascular smooth muscle cells].

    PubMed

    Yang, Guang; Zhang, Min-zhou; Jiang, Wei

    2005-10-01

    This paper sums up some studies in the last decade regarding the inhibitory effects of traditional Chinese herbs on growth factor induced proliferation of vascular smooth muscle cell (VSMC) via directly measuring the mRNA expression of its growth factors and the related receptors by electron microscope, immunohistochemistry, blot and hybridization in situ.

  15. Exogenous nerve growth factor protects the hypoglossal nerve against crush injury

    PubMed Central

    Fan, Li-yuan; Wang, Zhong-chao; Wang, Pin; Lan, Yu-yan; Tu, Ling

    2015-01-01

    Studies have shown that sensory nerve damage can activate the p38 mitogen-activated protein kinase (MAPK) pathway, but whether the same type of nerve injury after exercise activates the p38MAPK pathway remains unclear. Several studies have demonstrated that nerve growth factor may play a role in the repair process after peripheral nerve injury, but there has been little research focusing on the hypoglossal nerve injury and repair. In this study, we designed and established rat models of hypoglossal nerve crush injury and gave intraperitoneal injections of exogenous nerve growth factor to rats for 14 days. p38MAPK activity in the damaged neurons was increased following hypoglossal nerve crush injury; exogenous nerve growth factor inhibited this increase in acitivity and increased the survival rate of motor neurons within the hypoglossal nucleus. Under transmission electron microscopy, we found that the injection of nerve growth factor contributed to the restoration of the morphology of hypoglossal nerve after crush injury. Our experimental findings indicate that exogenous nerve growth factor can protect damaged neurons and promote hypoglossal nerve regeneration following hypoglossal nerve crush injury. PMID:26889186

  16. Hyaluronan modulates growth factor induced mammary gland branching in a size dependent manner.

    PubMed

    Tolg, Cornelia; Yuan, Han; Flynn, Sarah M; Basu, Kaustuv; Ma, Jenny; Tse, Kenneth Chor Kin; Kowalska, Beatrice; Vulkanesku, Diana; Cowman, Mary K; McCarthy, James B; Turley, Eva A

    2017-11-01

    Mammary gland morphogenesis begins during fetal development but expansion of the mammary tree occurs postnatally in response to hormones, growth factors and extracellular matrix. Hyaluronan (HA) is an extracellular matrix polysaccharide that has been shown to modulate growth factor-induced branching in culture. Neither the physiological relevance of HA to mammary gland morphogenesis nor the role that HA receptors play in these responses are currently well understood. We show that HA synthase (HAS2) is expressed in both ductal epithelia and stromal cells but HA primarily accumulates in the stroma. HA accumulation and expression of the HA receptors CD44 and RHAMM are highest during gestation when gland remodeling, lateral branch infilling and lobulo-alveoli formation is active. Molecular weight analyses show that approximately 98% of HA at all stages of morphogenesis is >300kDa. Low levels of 7-114kDa HA fragments are also detected and in particular the accumulation of 7-21kDa HA fragments are significantly higher during gestation than other morphogenetic stages (p<0.05). Using these in vivo results as a guide, in culture analyses of mammary epithelial cell lines (EpH4 and NMuMG) were performed to determine the roles of high molecular weight, 7-21kDa (10kDa MWavg) and HA receptors in EGF-induced branching morphogenesis. Results of these assays show that while HA synthesis is required for branching and 10kDa HA fragments strongly stimulate branching, the activity of HA decreases with increasing molecular weight and 500kDa HA strongly inhibits this morphogenetic process. The response to 10kDa HA requires RHAMM function and genetic deletion of RHAMM transiently blunts lateral branching in vivo. Collectively, these results reveal distinct roles for HA polymer size in modulating growth factor induced mammary gland branching and implicates these polymers in both the expansion and sculpting of the mammary tree during gestation. Copyright © 2017 Elsevier B.V. All rights

  17. Sciatic nerve regeneration using a nerve growth factor-containing fibrin glue membrane.

    PubMed

    Ma, Shengzhong; Peng, Changliang; Wu, Shiqing; Wu, Dongjin; Gao, Chunzheng

    2013-12-25

    Our previous findings confirmed that the nerve growth factor-containing fibrin glue membrane provides a good microenvironment for peripheral nerve regeneration; however, the precise mechanism remains unclear. p75 neurotrophin receptor (p75(NTR)) plays an important role in the regulation of peripheral nerve regeneration. We hypothesized that a nerve growth factor-containing fibrin glue membrane can promote neural regeneration by up-regulating p75(NTR) expression. In this study, we used a silicon nerve conduit to bridge a 15 mm-long sciatic nerve defect and injected a mixture of nerve growth factor and fibrin glue at the anastomotic site of the nerve conduit and the sciatic nerve. Through RT-PCR and western blot analysis, nerve growth factor-containing fibrin glue membrane significantly increased p75(NTR) mRNA and protein expression in the Schwann cells at the anastomotic site, in particular at 8 weeks after injection of the nerve growth factor/fibrin glue mixture. These results indicate that nerve growth factor-containing fibrin glue membrane can promote peripheral nerve regeneration by up-regulating p75(NTR) expression in Schwann cells.

  18. Interrelationship between growth factor-induced pH changes and intracellular Ca/sup 2 +/

    SciTech Connect

    Ives, H.E.; Daniel, T.O.

    1987-04-01

    Many mitogens cause rapid changes in intracellular pH and Ca/sup 2 +/. The authors studied the patterns of pH and Ca/sup 2 +/ changes after exposure of murine fibroblasts to platelet-derived growth factor (PDGF), bombesin, phorbol 12-myristate 13-acetate (PMA), and the vasoactive peptide bradykinin. Intracellular pH and Ca/sup 2 +/ were measured by using the fluorescent dyes 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein and fura-2. Three distinct patterns of intracellular pH change were observed. (i) PDGF and bombesin caused a rapid cytoplasmic acidification of 0.03 pH unit followed by a slower alkalinization of approx. = 0.11 pH unit above the resting pH of 6.88. (ii) PMA caused alkalinization without causing the early acidification. (iii) Bradykinin caused rapid acidification without the slower net alkalinization. All acidification responses were amiloride resistant. Patterns of intracellular Ca/sup 2 +/ response were also determined for each agent. In Ca/sup 2 +/-buffered cells, PDGF, bombesin, bradykinin, and ionomycin failed to induce cellular acidification, but alkalinization responses to PDGF, bombesin, and PMA persisted. They propose that the transient acidification seen with PDGF, bombesin, and other agents is the result of increased intracellular Ca/sup 2 +/. However, growth factor-induced alkalinization via the Na/sup +//H/sup +/ exchanger is independent of changes in Ca/sup 2 +/.

  19. Mo polyoxometalate nanoparticles inhibit tumor growth and vascular endothelial growth factor induced angiogenesis

    NASA Astrophysics Data System (ADS)

    Zheng, Wenjing; Yang, Licong; Liu, Ying; Qin, Xiuying; Zhou, Yanhui; Zhou, Yunshan; Liu, Jie

    2014-06-01

    Tumor growth depends on angiogenesis, which can furnish the oxygen and nutrients that proliferate tumor cells. Thus, blocking angiogenesis can be an effective strategy to inhibit tumor growth. In this work, three typical nanoparticles based on polyoxometalates (POMs) have been prepared; we investigated their capability as antitumor and anti-angiogenesis agents. We found that Mo POM nanoparticles, especially complex 3, inhibited the growth of human hepatocellular liver carcinoma cells (HepG2) through cellular reactive oxygen species levels’ elevation and mitochondrial membrane potential damage. Complex 3 also suppressed the proliferation, migration, and tube formation of endothelial cells in vitro and chicken chorioallantoic membrane development ex vivo. Furthermore, western blot analysis of cell signaling molecules indicated that Mo POMs blocked the vascular endothelial growth factor receptor 2-mediated ERK1/2 and AKT signaling pathways in endothelial cells. Using transmission electron microscopy, we demonstrated their cellular uptake and localization within the cytoplasm of HepG2 cells. These results indicate that, owing to the extraordinary physical and chemical properties, Mo POM nanoparticles can significantly inhibit tumor growth and angiogenesis, which makes them potential drug candidates in anticancer and anti-angiogenesis therapies.

  20. Mechanisms of nerve growth factor signaling in bone nociceptors and in an animal model of inflammatory bone pain.

    PubMed

    Nencini, Sara; Ringuet, Mitchell; Kim, Dong-Hyun; Chen, Yu-Jen; Greenhill, Claire; Ivanusic, Jason J

    2017-01-01

    Sequestration of nerve growth factor has been used successfully in the management of pain in animal models of bone disease and in human osteoarthritis. However, the mechanisms of nerve growth factor-induced bone pain and its role in modulating inflammatory bone pain remain to be determined. In this study, we show that nerve growth factor receptors (TrkA and p75) and some other nerve growth factor-signaling molecules (TRPV1 and Nav1.8, but not Nav1.9) are expressed in substantial proportions of rat bone nociceptors. We demonstrate that nerve growth factor injected directly into rat tibia rapidly activates and sensitizes bone nociceptors and produces acute behavioral responses with a similar time course. The nerve growth factor-induced changes in the activity and sensitivity of bone nociceptors we report are dependent on signaling through the TrkA receptor, but are not affected by mast cell stabilization. We failed to show evidence for longer term changes in expression of TrkA, TRPV1, Nav1.8 or Nav1.9 in the soma of bone nociceptors in a rat model of inflammatory bone pain. Thus, retrograde transport of NGF/TrkA and increased expression of some of the common nerve growth factor signaling molecules do not appear to be important for the maintenance of inflammatory bone pain. The findings are relevant to understand the basis of nerve growth factor sequestration and other therapies directed at nerve growth factor signaling, in managing pain in bone disease.

  1. Nerve Growth Factor and Diabetic Neuropathy

    PubMed Central

    Vinik, Aaron

    2003-01-01

    Neuropathy is one of the most debilitating complications of both type 1 and type 2 diabetes, with estimates of prevalence between 50–90% depending on the means of detection. Diabetic neuropathies are heterogeneous and there is variable involvement of large myelinated fibers and small, thinly myelinated fibers. Many of the neuronal abnormalities in diabetes can be duplicated by experimental depletion of specific neurotrophic factors, their receptors or their binding proteins. In experimental models of diabetes there is a reduction in the availability of these growth factors, which may be a consequence of metabolic abnormalities, or may be independent of glycemic control. These neurotrophic factors are required for the maintenance of the neurons, the ability to resist apoptosis and regenerative capacity. The best studied of the neurotrophic factors is nerve growth factor (NGF) and the related members of the neurotrophin family of peptides. There is increasing evidence that there is a deficiency of NGF in diabetes, as well as the dependent neuropeptides substance P (SP) and calcitonin gene-related peptide (CGRP) that may also contribute to the clinical symptoms resulting from small fiber dysfunction. Similarly, NT3 appears to be important for large fiber and IGFs for autonomic neuropathy. Whether the observed growth factor deficiencies are due to decreased synthesis, or functional, e.g. an inability to bind to their receptor, and/or abnormalities in nerve transport and processing, remains to be established. Although early studies in humans on the role of neurotrophic factors as a therapy for diabetic neuropathy have been unsuccessful, newer agents and the possibilities uncovered by further studies should fuel clinical trials for several generations. It seems reasonable to anticipate that neurotrophic factor therapy, specifically targeted at different nerve fiber populations, might enter the therapeutic armamentarium. PMID:14668049

  2. Nerve growth factor: neurotrophin or cytokine?

    PubMed

    Bonini, S; Rasi, G; Bracci-Laudiero, M L; Procoli, A; Aloe, L

    2003-06-01

    Nerve growth factor (NGF) is a neutrophin exerting an important role in the development and functions of the central and peripheral nervous system. However, it has recently been documented that several immune cells - such as mast cells, lymphocytes and eosinophils - produce, store and release NGF. Moreover, NGF high and low affinity receptors are widely expressed in the immune system, thus indicating the potential of responding to this neurotrophin through an autocrine mechanism. In fact, NGF influences development differentiation, chemotaxis and mediator release of inflammatory cells as well as fibroblast activation through a complex network influenced by other pro-inflammatory cytokines. Finally, NGF is increased in biological fluids of several allergic, immune and inflammatory diseases. Data reviewed suggest, therefore, that NGF might also be viewed as a (Th2?) cytokine with a modulatory role in allergic inflammation and tissue remodeling. Copyright 2003 S. Karger AG, Basel

  3. Phosphorylated endothelial nitric oxide synthase mediates vascular endothelial growth factor-induced penile erection.

    PubMed

    Musicki, Biljana; Palese, Michael A; Crone, Julie K; Burnett, Arthur L

    2004-02-01

    The objective of the present study was to evaluate whether vascular endothelial growth factor (VEGF)-induced penile erection is mediated by activation of endothelial nitric oxide synthase (eNOS) through its phosphorylation. We assessed the role of constitutively activated eNOS in VEGF-induced penile erection using wild-type (WT) and eNOS-knockout (eNOS(-/-)) mice with and without vasculogenic erectile dysfunction. Adult WT and eNOS(-/-) mice were subjected to sham operation or bilateral castration to induce vasculogenic erectile dysfunction. At the time of surgery, animals were injected intracavernosally with a replication-deficient adenovirus expressing human VEGF145 (10(9) particle units) or with empty virus (Ad.Null). After 7 days, erectile function was assessed in response to cavernous nerve electrical stimulation. Total and phosphorylated protein kinase B (Akt) as well as total and phosphorylated eNOS were quantitatively assessed in mice penes using Western immunoblot and immunohistochemistry. In intact WT mice, VEGF145 significantly increased erectile responses, and in WT mice after castration, it completely recovered penile erection. However, VEGF145 failed to increase erectile responses in intact eNOS(-/-) mice and only partially recovered erectile function in castrated eNOS(-/-) mice. In addition, VEGF145 significantly increased phosphorylation of eNOS at Serine 1177 by approximately 2-fold in penes of both intact and castrated WT mice. The data provide a molecular explanation for VEGF stimulatory effect on penile erection, which involves phosphorylated eNOS (Serine 1177) mediation.

  4. Expression of nerve growth factor and its receptor in distracted tibial nerve after limb lengthening.

    PubMed

    Shao, Heng; Shu, Hengsheng; Wang, Chunmei; Yuan, Wu; Li, Yunsheng

    2013-02-01

    Despite many experimental and clinical studies conducted on distraction osteogenesis (DO) in the past decade, changes in the surrounding tissues that occur after the procedure remains poorly understood. To study the biochemical changes of recovery in nerve tissues upon DO-induced nerve injury, we prepared a rabbit model of tibia lengthening to observe the expression pattern of nerve growth factor (NGF) and low-affinity NGF receptor (p75NGFR) in the distracted tibial nerve. The distracted tibial nerve was harvested at various time points during the consolidation period of new bone formation and immunohistochemical staining was performed to detect the expression of NGF and p75NGFR. The expression levels of NGF and p75NGFR were found to be different at various times after DO. The changes in expression of these two cellular factors show similar tendencies with significantly elevated expression in Schwann cells at 7 and 14 days after distraction, but low or undetectable levels of expression at 0, 28, and 56 days. These results suggest that NGF and p75NGFR may play important roles in the adaptive process of the distracted nerve. NGF and p75NGFR are autocrine growth factors present in the distracted nerve during the early consolidation period. NGF interacts with p75NGFR to promote damage repair and reconstruction of nerves. Together, this study furthers the understanding of the relative mechanisms of nerve repair, as well as provides a further basis for the clinical application of neurotrophins.

  5. Nerve growth factor actions on the brain

    SciTech Connect

    Martinez, H.J.

    1989-01-01

    We examined the effect of the trophic protein, nerve growth factor (NGF), on cultures of fetal rat neostriatum and basal forebrain-medial septal area (BF-MS) to define its role in brain development. Treatment of cultures with NGF resulted in an increase in the specific activity of the cholinergic enzyme choline acetyltransferase (CAT) in both brain areas. CAT was immunocytochemically localized to neurons. In the BF-MS, NGF treatment elicited a marked increase in staining intensity and an apparent increase in the number of CAT-positive neurons. Moreover, treatment of BF-MS cultures with NGF increased the activity of acetylcholinesterase, suggesting that the cholinergic neuron as a whole was affected. To begin defining mechanisms of action of NGF in the BF-MS, we detected NGF receptors by two independent methods. Receptors were localized to two different cellular populations: neuron-like cells, and non-neuron-like cells. Dissociation studies with ({sup 125}I)NGF suggested that high affinity receptors were localized to the neuron-like population. Only low-affinity receptors were localized to the non-neuron-like cells. Moreover, employing combined immunocytochemistry and ({sup 125}I)NGF autoradiography, we detected a subpopulation of CAT-containing neutrons that exhibited high-affinity binding. Unexpectedly, a gamma-aminobutyric acid (GABA)-containing cell group also expressed high affinity binding. However, only subsets of cholinergic or GABA neurons expressed high-affinity biding, suggesting that these transmitter populations are composed of differentially response subpopulations.

  6. Nerve growth factor enhances sleep in rabbits.

    PubMed

    Takahashi, S; Krueger, J M

    1999-04-02

    Nerve growth factor (NGF) elicits rapid-eye-movement sleep (REMS) in cats. Removal of NGF receptor-positive cholinergic basal forebrain neurons inhibits REMS in rats. The aim of the present study was to determine the effects of NGF on sleep and brain temperature (Tbr) in rabbits. Male rabbits were implanted with electroencephalograph (EEG) electrodes, a brain thermistor and an intraventricular (i.c.v.) guide cannula. Rabbits received human beta-NGF i.c.v. (0.01, 0.1, 1.0 or 10 microg] and on a separate day, 25 microl pyrogen-free saline i.c.v. as control. EEG and Tbr were recorded for 23 h after injections. The highest two doses of NGF increased both non-REMS and REMS across the 23-h recording period. REMS was enhanced dose-dependently. Tbr was not affected by any dose of NGF. These results suggest that NGF is involved in both REMS and non-REMS regulation.

  7. Nerve growth factor promotes human hemopoietic colony growth and differentiation.

    PubMed Central

    Matsuda, H; Coughlin, M D; Bienenstock, J; Denburg, J A

    1988-01-01

    Nerve growth factor (NGF) is a neurotropic polypeptide necessary for the survival and growth of some central neurons, as well as sensory afferent and sympathetic neurons. Much is now known of the structural and functional characteristics of NGF, whose gene has recently been cloned. Since it is synthesized in largest amounts by the male mouse submandibular gland, its role exclusively in nerve growth is questionable. NGF also causes histamine release from rat peritoneal mast cells in vitro, and we have shown elsewhere that it causes significant, dose-dependent, generalized mast cell proliferation in the rat in vivo when administered neonatally. Our experiments now indicate that NGF causes a significant stimulation of granulocyte colonies grown from human peripheral blood in standard hemopoietic methylcellulose assays. Further, NGF appears to act in a relatively selective fashion to induce the differentiation of eosinophils and basophils/mast cells. Depletion experiments show that the NGF effect may be T-cell dependent and that NGF augments the colony-stimulating effect of supernatants from the leukemic T-cell (Mo) line. The hemopoietic activity of NGF is blocked by polyclonal and monoclonal antibodies to NGF. We conclude that NGF may indirectly act as a local growth factor in tissues other than those of the nervous system by causing T cells to synthesize or secrete molecules with colony-stimulating activity. In view of the synthesis of NGF in tissue injury, the involvement of basophils/mast cells and eosinophils in allergic and other inflammatory processes, and the association of mast cells with fibrosis and tissue repair, we postulate that NGF plays an important biological role in a variety of repair processes. PMID:3413109

  8. RAPID COMMUNICATION: Nerve growth factor influences cleavage rate and embryo development in sheep.

    PubMed

    Crispo, M; Dos Santos-Neto, P C; Vilariño, M; Mulet, A P; de León, A; Barbeito, L; Menchaca, A

    2016-10-01

    Recent information about Nerve growth factor (NGF), a protein traditionally associated to the nervous system that regulates survival and maturation of developing neurons, suggests that it may exert action also on different levels in the reproductive system. The aim of this study was to evaluate the effect of NGF added during in vitro oocyte maturation, fertilization or in vitro embryo development in sheep. Nerve growth factor was supplemented to the culture medium at 0, 100, or 1,000 ng/mL, during either in vitro maturation (Exp. 1), in vitro fertilization (Exp. 2), or in vitro culture (Exp. 3). In addition, NGF mRNA expression was determined in cumulus cells and oocytes. Nerve growth factor induced early cleavage when added during oocyte maturation or fertilization, improved embryo development when added during fertilization, and had no significant effect when added during embryo culture. In general, the effect was more evident with 100 rather than 1,000 ng/mL (P < 0.05). Expression of endogenous NGF was not detected in oocytes, and increased in cumulus cells when 1,000 ng/mL of NGF was added during fertilization, but not during maturation and embryo culture. In conclusion, the addition of NGF during oocyte maturation and fertilization affects in vitro cleavage and embryo development in sheep. We suggest a possible effect of this growth factor on oocyte maturation and mainly on the fertilization process.

  9. Let-7 microRNAs regenerate peripheral nerve regeneration by targeting nerve growth factor.

    PubMed

    Li, Shiying; Wang, Xinghui; Gu, Yun; Chen, Chu; Wang, Yaxian; Liu, Jie; Hu, Wen; Yu, Bin; Wang, Yongjun; Ding, Fei; Liu, Yan; Gu, Xiaosong

    2015-03-01

    Peripheral nerve injury is a common clinical problem. Nerve growth factor (NGF) promotes peripheral nerve regeneration, but its clinical applications are limited by several constraints. In this study, we found that the time-dependent expression profiles of eight let-7 family members in the injured nerve after sciatic nerve injury were roughly similar to each other. Let-7 microRNAs (miRNAs) significantly reduced cell proliferation and migration of primary Schwann cells (SCs) by directly targeting NGF and suppressing its protein translation. Following sciatic nerve injury, the temporal change in let-7 miRNA expression was negatively correlated with that in NGF expression. Inhibition of let-7 miRNAs increased NGF secretion by primary cultured SCs and enhanced axonal outgrowth from a coculture of primary SCs and dorsal root gangalion neurons. In vivo tests indicated that let-7 inhibition promoted SCs migration and axon outgrowth within a regenerative microenvironment. In addition, the inhibitory effect of let-7 miRNAs on SCs apoptosis might serve as an early stress response to nerve injury, but this effect seemed to be not mediated through a NGF-dependent pathway. Collectively, our results provide a new insight into let-7 miRNA regulation of peripheral nerve regeneration and suggest a potential therapy for repair of peripheral nerve injury.

  10. Vascular endothelial growth factor mediates corneal nerve repair.

    PubMed

    Yu, Charles Q; Zhang, Min; Matis, Krisztina I; Kim, Charles; Rosenblatt, Mark I

    2008-09-01

    To examine the expression of vascular endothelial growth factor (VEGF) and its receptors in the cornea and the trigeminal ganglion and to characterize the role of VEGF in mediating corneal nerve repair. Regeneration of the corneal subbasal nerve plexus after epithelial debridement was measured. The expression of VEGF and its receptors was examined in the trigeminal ganglia and in the cornea by RT-PCR, immunohistochemistry, and Western blotting. VEGF-mediated nerve growth was measured in a trigeminal ganglia explant assay. Anti-VEGF neutralizing antibody was used to examine the VEGF-dependent growth of neurons in vitro and regeneration of the corneal nerves in vivo. After two distinct patterns of nerve regeneration, the subbasal nerves recovered to 65% of the preinjury density after 28 days. RT-PCR demonstrated gene expression of VEGF and VEGF receptors in the trigeminal ganglia. Immunohistochemistry showed staining for VEGF and its receptors in the trigeminal ganglia and for VEGFR1, VEGFR2, and neuropilin (NRP)-1 in the cornea. Western blot confirmed these results. In vitro, VEGF promoted the growth of explanted trigeminal ganglia by 91%. Blockage of VEGF signaling with anti-VEGF antibody reduced the growth of cultured neurons by 17% and the regeneration of subbasal neurons by 23%. In addition to providing new information on the regeneration of murine corneal nerves, this study presents evidence that VEGF signaling influences the repair of corneal nerves by demonstrating that VEGF and VEGF receptors are present in the trigeminal ganglia and that abrogation of VEGF signaling reduces nerve growth in vitro and in vivo.

  11. Obesity and Postmenopausal Breast Cancer Risk: Determining the Role of Growth Factor-Induced Aromatase Expression

    DTIC Science & Technology

    2014-03-01

    levels of insulin, insulin-like growth factor 1 (IGF-1), and leptin . These all have significant tumorigenic effects in the breast and are known to...role of obesity-associated growth factor and leptin signaling on aromatase expression and activity, we will also add inhibitors to these different

  12. Peripheral Nerve Regeneration Strategies: Electrically Stimulating Polymer Based Nerve Growth Conduits

    PubMed Central

    Anderson, Matthew; Shelke, Namdev B.; Manoukian, Ohan S.; Yu, Xiaojun; McCullough, Louise D.; Kumbar, Sangamesh G.

    2017-01-01

    Treatment of large peripheral nerve damages ranges from the use of an autologous nerve graft to a synthetic nerve growth conduit. Biological grafts, in spite of many merits, show several limitations in terms of availability and donor site morbidity, and outcomes are suboptimal due to fascicle mismatch, scarring, and fibrosis. Tissue engineered nerve graft substitutes utilize polymeric conduits in conjunction with cues both chemical and physical, cells alone and or in combination. The chemical and physical cues delivered through polymeric conduits play an important role and drive tissue regeneration. Electrical stimulation (ES) has been applied toward the repair and regeneration of various tissues such as muscle, tendon, nerve, and articular tissue both in laboratory and clinical settings. The underlying mechanisms that regulate cellular activities such as cell adhesion, proliferation, cell migration, protein production, and tissue regeneration following ES is not fully understood. Polymeric constructs that can carry the electrical stimulation along the length of the scaffold have been developed and characterized for possible nerve regeneration applications. We discuss the use of electrically conductive polymers and associated cell interaction, biocompatibility, tissue regeneration, and recent basic research for nerve regeneration. In conclusion, a multifunctional combinatorial device comprised of biomaterial, structural, functional, cellular, and molecular aspects may be the best way forward for effective peripheral nerve regeneration. PMID:27278739

  13. SOXC Transcription Factors Induce Cartilage Growth Plate Formation in Mouse Embryos by Promoting Noncanonical WNT Signaling.

    PubMed

    Kato, Kenji; Bhattaram, Pallavi; Penzo-Méndez, Alfredo; Gadi, Abhilash; Lefebvre, Véronique

    2015-09-01

    Growth plates are specialized cartilage structures that ensure the elongation of most skeletal primordia during vertebrate development. They are made by chondrocytes that proliferate in longitudinal columns and then progress in a staggered manner towards prehypertrophic, hypertrophic and terminal maturation. Complex molecular networks control the formation and activity of growth plates, but remain incompletely understood. We investigated here the importance of the SoxC genes, which encode the SOX4, SOX11 and SOX12 transcription factors, in growth plates. We show that the three genes are expressed robustly in perichondrocytes and weakly in growth plate chondrocytes. SoxC(Prx1Cre) mice, which deleted SoxC genes in limb bud skeletogenic mesenchyme, were born with tiny appendicular cartilage primordia because of failure to form growth plates. In contrast, SoxC(Col2Cre) and SoxC(ATC) mice, which deleted SoxC genes primarily in chondrocytes, were born with mild dwarfism and fair growth plates. Chondrocytes in the latter mutants matured normally, but formed irregular columns, proliferated slowly and died ectopically. Asymmetric distribution of VANGL2 was defective in both SoxC(Prx1Cre) and SoxC(ATC) chondrocytes, indicating impairment of planar cell polarity, a noncanonical WNT signaling pathway that controls growth plate chondrocyte alignment, proliferation and survival. Accordingly, SoxC genes were necessary in perichondrocytes for expression of Wnt5a, which encodes a noncanonical WNT ligand required for growth plate formation, and in chondrocytes and perichondrocytes for expression of Fzd3 and Csnk1e, which encode a WNT receptor and casein kinase-1 subunit mediating planar cell polarity, respectively. Reflecting the differential strengths of the SOXC protein transactivation domains, SOX11 was more powerful than SOX4, and SOX12 interfered with the activity of SOX4 and SOX11. Altogether, these findings provide novel insights into the molecular regulation of skeletal

  14. Protein kinase A regulates 3-phosphatidylinositide dynamics during platelet-derived growth factor-induced membrane ruffling and chemotaxis.

    PubMed

    Deming, Paula B; Campbell, Shirley L; Baldor, Linda C; Howe, Alan K

    2008-12-12

    Spatial regulation of the cAMP-dependent protein kinase (PKA) is required for chemotaxis in fibroblasts; however, the mechanism(s) by which PKA regulates the cell migration machinery remain largely unknown. Here we report that one function of PKA during platelet-derived growth factor (PDGF)-induced chemotaxis was to promote membrane ruffling by regulating phosphatidylinositol 3,4,5-trisphosphate (PIP(3)) dynamics. Inhibition of PKA activity dramatically altered membrane dynamics and attenuated formation of peripheral membrane ruffles in response to PDGF. PKA inhibition also significantly decreased the number and size of PIP(3)-rich membrane ruffles in response to uniform stimulation and to gradients of PDGF. This ruffling defect was quantified using a newly developed method, based on computer vision edge-detection algorithms. PKA inhibition caused a marked attenuation in the bulk accumulation of PIP(3) following PDGF stimulation, without effects on PI3-kinase (PI3K) activity. The deficits in PIP(3) dynamics correlated with a significant inhibition of growth factor-induced membrane recruitment of endogenous Akt and Rac activation in PKA-inhibited cells. Simultaneous inhibition of PKA and Rac had an additive inhibitory effect on growth factor-induced ruffling dynamics. Conversely, the expression of a constitutively active Rac allele was able to rescue the defect in membrane ruffling and restore the localization of a fluorescent PIP(3) marker to membrane ruffles in PKA-inhibited cells, even in the absence of PI3K activity. These data demonstrate that, like Rac, PKA contributes to PIP(3) and membrane dynamics independently of direct regulation of PI3K activity and suggest that modulation of PIP(3)/3-phosphatidylinositol (3-PI) lipids represents a major target for PKA in the regulation of PDGF-induced chemotactic events.

  15. Opposing functions of TFII-I spliced isoforms in growth factor-induced gene expression.

    PubMed

    Hakre, Shweta; Tussie-Luna, María Isabel; Ashworth, Todd; Novina, Carl D; Settleman, Jeffrey; Sharp, Phillip A; Roy, Ananda L

    2006-10-20

    Multifunctional transcription factor TFII-I has two spliced isoforms (Delta and beta) in murine fibroblasts. Here we show that these isoforms have distinct subcellular localization and mutually exclusive transcription functions in the context of growth factor signaling. In the absence of signaling, TFII-Ibeta is nuclear and recruited to the c-fos promoter in vivo. But upon growth factor stimulation, the promoter recruitment is abolished and it is exported out of the nucleus. Moreover, isoform-specific silencing of TFII-Ibeta results in transcriptional activation of the c-fos gene. In contrast, TFII-IDelta is largely cytoplasmic in the resting state but translocates to the nucleus upon growth factor signaling, undergoes signal-induced recruitment to the same site on the c-fos promoter, and activates the gene. Importantly, activated TFII-IDelta interacts with Erk1/2 (MAPK) kinase in the cell cytoplasm and imports the Erk1/2 to the nucleus, thereby transducing growth factor signaling. Our results identify a unique growth factor signaling pathway controlled by opposing activities of two TFII-I spliced isoforms.

  16. Rapamycin promotes Schwann cell migration and nerve growth factor secretion

    PubMed Central

    Liu, Fang; Zhang, Haiwei; Zhang, Kaiming; Wang, Xinyu; Li, Shipu; Yin, Yixia

    2014-01-01

    Rapamycin, similar to FK506, can promote neural regeneration in vitro. We assumed that the mechanisms of action of rapamycin and FK506 in promoting peripheral nerve regeneration were similar. This study compared the effects of different concentrations of rapamycin and FK506 on Schwann cells and investigated effects and mechanisms of rapamycin on improving peripheral nerve regeneration. Results demonstrated that the lowest rapamycin concentration (1.53 nmol/L) more significantly promoted Schwann cell migration than the highest FK506 concentration (100μmol/L). Rapamycin promoted the secretion of nerve growth factors and upregulated growth-associated protein 43 expression in Schwann cells, but did not significantly affect Schwann cell proliferation. Therefore, rapamycin has potential application in peripheral nerve regeneration therapy. PMID:25206862

  17. 2D protrusion but not motility predicts growth factor-induced cancer cell migration in 3D collagen.

    PubMed

    Meyer, Aaron S; Hughes-Alford, Shannon K; Kay, Jennifer E; Castillo, Amalchi; Wells, Alan; Gertler, Frank B; Lauffenburger, Douglas A

    2012-06-11

    Growth factor-induced migration is a critical step in the dissemination and metastasis of solid tumors. Although differences in properties characterizing cell migration on two-dimensional (2D) substrata versus within three-dimensional (3D) matrices have been noted for particular growth factor stimuli, the 2D approach remains in more common use as an efficient surrogate, especially for high-throughput experiments. We therefore were motivated to investigate which migration properties measured in various 2D assays might be reflective of 3D migratory behavioral responses. We used human triple-negative breast cancer lines stimulated by a panel of receptor tyrosine kinase ligands relevant to mammary carcinoma progression. Whereas 2D migration properties did not correlate well with 3D behavior across multiple growth factors, we found that increased membrane protrusion elicited by growth factor stimulation did relate robustly to enhanced 3D migration properties of the MDA-MB-231 and MDA-MB-157 lines. Interestingly, we observed this to be a more reliable relationship than cognate receptor expression or activation levels across these and two additional mammary tumor lines.

  18. Facilitation of facial nerve regeneration using chitosan-β-glycerophosphate-nerve growth factor hydrogel.

    PubMed

    Chao, Xiuhua; Xu, Lei; Li, Jianfeng; Han, Yuechen; Li, Xiaofei; Mao, YanYan; Shang, Haiqiong; Fan, Zhaomin; Wang, Haibo

    2016-06-01

    Conclusion C/GP hydrogel was demonstrated to be an ideal drug delivery vehicle and scaffold in the vein conduit. Combined use autologous vein and NGF continuously delivered by C/GP-NGF hydrogel can improve the recovery of facial nerve defects. Objective This study investigated the effects of chitosan-β-glycerophosphate-nerve growth factor (C/GP-NGF) hydrogel combined with autologous vein conduit on the recovery of damaged facial nerve in a rat model. Methods A 5 mm gap in the buccal branch of a rat facial nerve was reconstructed with an autologous vein. Next, C/GP-NGF hydrogel was injected into the vein conduit. In negative control groups, NGF solution or phosphate-buffered saline (PBS) was injected into the vein conduits, respectively. Autologous implantation was used as a positive control group. Vibrissae movement, electrophysiological assessment, and morphological analysis of regenerated nerves were performed to assess nerve regeneration. Results NGF continuously released from C/GP-NGF hydrogel in vitro. The recovery rate of vibrissae movement and the compound muscle action potentials of regenerated facial nerve in the C/GP-NGF group were similar to those in the Auto group, and significantly better than those in the NGF group. Furthermore, larger regenerated axons and thicker myelin sheaths were obtained in the C/GP-NGF group than those in the NGF group.

  19. The role of EGF receptor transmodulation in embryonal carcinoma-derived growth factor-induced mitogenesis.

    PubMed Central

    Heath, J K; Mahadevan, L; Foulkes, J G

    1986-01-01

    Exposure of quiescent 10T1/2 fibroblast cells to embryonal carcinoma-derived growth factor (ECDGF) results in a rapid temperature and ECDGF concentration-dependent inhibition of [125I]EGF binding to the epidermal growth factor (EGF) receptor (transmodulation). ECDGF predominantly inhibits the association of [125I]EGF with a high affinity subclass of EGF receptors, and induces increased phosphorylation of the EGF receptor on serine and threonine residues. No mitogenic effect of EGF can be detected in the presence of ECDGF concentrations which induce maximal EGF receptor transmodulation. ECDGF-induced EGF receptor transmodulation is sensitive to phorbol ester-induced desensitization whereas ECDGF-induced DNA synthesis is unaffected by prolonged pre-treatment with biologically active phorbol ester. These findings suggest that EGF receptor transmodulation is not essential for ECDGF mitogenicity but may inhibit EGF-induced DNA synthesis. Images Fig. 5. PMID:3489616

  20. Mediation of growth factor induced DNA synthesis and calcium mobilization by Gq and Gi2

    PubMed Central

    1993-01-01

    A newly identified subclass of the heterotrimeric GTP binding regulatory protein family, Gq, has been found to be expressed in a diverse range of cell types. We investigated the potential role of this protein in growth factor signal transduction pathways and its potential relationship to the function of other G alpha subclasses. Recent biochemical studies have suggested that Gq regulates the beta 1 isozyme of phospholipase C (PLC beta 1), an effector for some growth factors. By microinjection of inhibitory antibodies specific to distinct G alpha subunits into living cells, we have determined that G alpha q transduces bradykinin- and thrombin-stimulated intracellular calcium transients which are likely to be mediated by PLC beta 1. Moreover, we found that G alpha q function is required for the mitogenic action of both of these growth factors. These results indicate that both thrombin and bradykinin utilize Gq to couple to increases in intracellular calcium, and that Gq is a necessary component of the mitogenic action of these factors. While microinjection of antibodies against G alpha i2 did not abolish calcium transients stimulated by either of these factors, such microinjection prevented DNA synthesis in response to thrombin but not to bradykinin. These data suggest that thrombin- induced mitogenesis requires both Gq and Gi2, whereas bradykinin needs only the former. Thus, different growth factors operating upon the same cell type use overlapping yet distinct sets of G alpha subtypes in mitogenic signal transduction pathways. The direct identification of the coupling of both a pertussis toxin sensitive and insensitive G protein subtype in the mitogenic pathways utilized by thrombin offers an in vivo biochemical clarification of previous results obtained by pharmacologic studies. PMID:8458876

  1. Keratinocyte growth factor induces matrix metalloproteinase-9 expression and correlates with venous invasion in pancreatic cancer

    PubMed Central

    CHO, KAZUMITSU; MATSUDA, YOKO; UEDA, JUNJI; UCHIDA, EIJI; NAITO, ZENYA; ISHIWATA, TOSHIYUKI

    2012-01-01

    Keratinocyte growth factor (KGF), also known as fibroblast growth factor-7, and KGF receptor (KGFR) play important roles in the growth of epithelial cells and are overexpressed in a variety of malignant epithelial tumors, including pancreatic ductal adenocarcinoma (PDAC). We previously reported that co-expression of KGF and KGFR in PDAC is associated with venous invasion, enhanced vascular endothelial growth factor A expression and poor prognosis. Matrix metalloproteinase-9 (MMP-9) is known to participate in the degradation of type IV collagen, which is a primary component of extracellular matrices in the vascular basement membrane. In the present study, we examined the expression and roles of KGF, KGFR and MMP-9 in human PDAC cell lines and tissues. Quantitative real-time polymerase chain reaction analysis demonstrated the expression of MMP-9 mRNA in all eight PDAC cell lines. KGF, KGFR and MMP-9 were, respectively, expressed in 27 (43%), 23 (37%) and 35 (56%) of 63 patients. Each expression of KGF, KGFR or MMP-9 correlated positively with venous invasion. Furthermore, expression of KGF or MMP-9 correlated positively with liver metastasis. KGF-positive cases exhibited shorter survival than KGF-negative cases, while KGFR and MMP-9 expression were unrelated to prognosis. Administration of recombinant human KGF increased MMP-9 expression in PDAC cells, while transient transfection with short hairpin RNAs targeting KGF transcripts reduced MMP-9 expression in PDAC cells. Moreover, recombinant human KGF significantly enhanced migration and invasion of PDAC cells. These findings suggest that KGF and KGFR promote venous invasion via MMP-9 in PDAC, and closely correlate with liver metastasis. The KGF/KGFR pathway may be a critical therapeutic target for PDAC metastasis. PMID:22159401

  2. Keratinocyte growth factor induces proliferation of hepatocytes and epithelial cells throughout the rat gastrointestinal tract.

    PubMed

    Housley, R M; Morris, C F; Boyle, W; Ring, B; Biltz, R; Tarpley, J E; Aukerman, S L; Devine, P L; Whitehead, R H; Pierce, G F

    1994-11-01

    Keratinocyte growth factor (KGF), a member of the fibroblast growth factor (FGF) family, was identified as a specific keratinocyte mitogen after isolation from a lung fibroblast line. Recently, recombinant (r)KGF was found to influence proliferation and differentiation patterns of multiple epithelial cell lineages within skin, lung, and the reproductive tract. In the present study, we designed experiments to identify additional target tissues, and focused on the rat gastrointestinal (GI) system, since a putative receptor, K-sam, was originally identified in a gastric carcinoma. Expression of KGF receptor and KGF mRNA was detected within the entire GI tract, suggesting the gut both synthesized and responded to KGF. Therefore, rKGF was administered to adult rats and was found to induce markedly increased proliferation of epithelial cells from the foregut to the colon, and of hepatocytes, one day after systemic treatment. Daily treatment resulted in the marked selective induction of mucin-producing cell lineages throughout the GI tract in a dose-dependent fashion. Other cell lineages were either unaffected (e.g., Paneth cells), or relatively decreased (e.g., parietal cells, enterocytes) in rKGF-treated rats. The direct effect of rKGF was confirmed by demonstrating markedly increased carcinoembryonic antigen production in a human colon carcinoma cell line, LIM1899. Serum levels of albumin were specifically and significantly elevated after daily treatment. These results demonstrate rKGF can induce epithelial cell activation throughout the GI tract and liver. Further, endogenous KGF may be a normal paracrine mediator of growth within the gut.

  3. Platelet-derived growth factor induces phosphorylation of a 64-kDa nuclear protein

    SciTech Connect

    Shawver, L.K.; Pierce, G.F.; Kawahara, R.S.; Deuel, T.F.

    1989-01-15

    The platelet-derived growth factor (PDGF) stimulated the phosphorylation of a nuclear protein of 64 kDa (pp64) in nuclei of nontransformed normal rat kidney (NRK) cells. Low levels of phosphorylation of pp64 were observed in nuclei of serum-starved NRK cells. Fetal calf serum (FCS), PDGF, and homodimeric v-sis and PDGF A-chain protein enhanced the incorporation of 32P into pp64 over 4-fold within 30 min and over 8-fold within 2 h of exposure of NRK cells to the growth factors. In contrast, constitutive phosphorylation of 32P-labeled pp64 in nuclei of NRK cells transformed by the simian sarcoma virus (SSV) was high and only minimally stimulated by PDGF and FCS. 32P-Labeled pp64 was isolated from nuclei of PDGF-stimulated nontransformed NRK cells; the 32P of pp64 was labile in 1 M KOH, and pp64 was not significantly recognized by anti-phosphotyrosine antisera, suggesting that the PDGF-induced phosphorylation of pp64 occurred on serine or on threonine residues. However, pp64 from SSV-transformed NRK cell nuclei was significantly stable to base hydrolysis and was immunoprecipitated with anti-phosphotyrosine antisera, suggesting that pp64 from SSV-transformed cell nuclei is phosphorylated also on tyrosine. FCS, PDGF, and PDGF A- and B-chain homodimers thus stimulate the rapid time-dependent phosphorylation of a 64-kDa nuclear protein shortly after stimulation of responsive cells. The growth factor-stimulated phosphorylation of pp64 and the constitutive high levels of pp64 phosphorylation in cells transformed by SSV suggest important roles for pp64 and perhaps regulated nuclear protein kinases and phosphatases in cell division and proliferation.

  4. Growth factors induce monocyte binding to vascular smooth muscle cells: implications for monocyte retention in atherosclerosis.

    PubMed

    Cai, Qiangjun; Lanting, Linda; Natarajan, Rama

    2004-09-01

    Adhesive interactions between monocytes and vascular smooth muscle cells (VSMC) may contribute to subendothelial monocyte-macrophage retention in atherosclerosis. We investigated the effects of angiotensin II (ANG II) and platelet-derived growth factor (PDGF)-BB on VSMC-monocyte interactions. Treatment of human aortic VSMC (HVSMC) with ANG II or PDGF-BB significantly increased binding to human monocytic THP-1 cells and to peripheral blood monocytes. This was inhibited by antibodies to monocyte beta(1)- and beta(2)-integrins. The binding was also attenuated by blocking VSMC arachidonic acid (AA) metabolism by inhibitors of 12/15-lipoxygenase (12/15-LO) or cyclooxygenase-2 (COX-2). Conversely, binding was enhanced by overexpression of 12/15-LO or COX-2. Direct treatment of HVSMC with AA or its metabolites also increased binding. Furthermore, VSMC derived from 12/15-LO knockout mice displayed reduced binding to mouse monocytic cells relative to genetic control mice. Using specific signal transduction inhibitors, we demonstrated the involvement of Src, phosphoinositide 3-kinase, and MAPKs in ANG II- or PDGF-BB-induced binding. Interestingly, after coculture with HVSMC, THP-1 cell surface expression of the scavenger receptor CD36 was increased. These results show for the first time that growth factors may play additional roles in atherosclerosis by increasing monocyte binding to VSMC via AA metabolism and key signaling pathways. This can lead to monocyte subendothelial retention, CD36 expression, and foam cell formation.

  5. [Sphingosine kinase regulates hepatocyte growth factor-induced migration of endothelial cells].

    PubMed

    Yi, Jun; Lu, Zhuao-Zhuang; Duan, Hai-Feng; Gai, Lu-Yue; Wang, Li-Sheng

    2006-05-01

    To elucidate the effect of sphingosine kinase (SPK) on the hepatocyte growth factor (HGF)-induced migration of endothelial cells. We constructed recombinant adenoviral vectors, which contain SPK gene and its mutant respectively. These adenoviral vectors were packaged and amplified in 293 cells. And intracellular SPK activity was assayed via measurement of [32]P radioisotope labeled S1P; the effect of SPK activation on HGF-induced migration of endothelial cell was observed by Transwell technique. Adenoviral mediated expression of SPK gene increased in ECV 304 cells intracellular SPK activity, which in turn enhanced the HGF-induced migration. Whereas these activities were blocked by the dominant negative SPK gene. These findings show that SPK activation plays important roles in the regulation of HGF-induced migration of endothelial cells.

  6. Sphingosine-1-phosphate mediates epidermal growth factor-induced muscle satellite cell activation.

    PubMed

    Nagata, Yosuke; Ohashi, Kazuya; Wada, Eiji; Yuasa, Yuki; Shiozuka, Masataka; Nonomura, Yoshiaki; Matsuda, Ryoichi

    2014-08-01

    Skeletal muscle can regenerate repeatedly due to the presence of resident stem cells, called satellite cells. Because satellite cells are usually quiescent, they must be activated before participating in muscle regeneration in response to stimuli such as injury, overloading, and stretch. Although satellite cell activation is a crucial step in muscle regeneration, little is known of the molecular mechanisms controlling this process. Recent work showed that the bioactive lipid sphingosine-1-phosphate (S1P) plays crucial roles in the activation, proliferation, and differentiation of muscle satellite cells. We investigated the role of growth factors in S1P-mediated satellite cell activation. We found that epidermal growth factor (EGF) in combination with insulin induced proliferation of quiescent undifferentiated mouse myoblast C2C12 cells, which are also known as reserve cells, in serum-free conditions. Sphingosine kinase activity increased when reserve cells were stimulated with EGF. Treatment of reserve cells with the D-erythro-N,N-dimethylsphingosine, Sphingosine Kinase Inhibitor, or siRNA duplexes specific for sphingosine kinase 1, suppressed EGF-induced C2C12 activation. We also present the evidence showing the S1P receptor S1P2 is involved in EGF-induced reserve cell activation. Moreover, we demonstrated a combination of insulin and EGF promoted activation of satellite cells on single myofibers in a manner dependent on SPHK and S1P2. Taken together, our observations show that EGF-induced satellite cell activation is mediated by S1P and its receptor.

  7. Sphingosine-1-phosphate mediates epidermal growth factor-induced muscle satellite cell activation

    SciTech Connect

    Nagata, Yosuke Ohashi, Kazuya; Wada, Eiji; Yuasa, Yuki; Shiozuka, Masataka; Nonomura, Yoshiaki; Matsuda, Ryoichi

    2014-08-01

    Skeletal muscle can regenerate repeatedly due to the presence of resident stem cells, called satellite cells. Because satellite cells are usually quiescent, they must be activated before participating in muscle regeneration in response to stimuli such as injury, overloading, and stretch. Although satellite cell activation is a crucial step in muscle regeneration, little is known of the molecular mechanisms controlling this process. Recent work showed that the bioactive lipid sphingosine-1-phosphate (S1P) plays crucial roles in the activation, proliferation, and differentiation of muscle satellite cells. We investigated the role of growth factors in S1P-mediated satellite cell activation. We found that epidermal growth factor (EGF) in combination with insulin induced proliferation of quiescent undifferentiated mouse myoblast C2C12 cells, which are also known as reserve cells, in serum-free conditions. Sphingosine kinase activity increased when reserve cells were stimulated with EGF. Treatment of reserve cells with the D-erythro-N,N-dimethylsphingosine, Sphingosine Kinase Inhibitor, or siRNA duplexes specific for sphingosine kinase 1, suppressed EGF-induced C2C12 activation. We also present the evidence showing the S1P receptor S1P2 is involved in EGF-induced reserve cell activation. Moreover, we demonstrated a combination of insulin and EGF promoted activation of satellite cells on single myofibers in a manner dependent on SPHK and S1P2. Taken together, our observations show that EGF-induced satellite cell activation is mediated by S1P and its receptor. - Highlights: • EGF in combination with insulin induces proliferation of quiescent C2C12 cells. • Sphingosine kinase activity increases when reserve cells are stimulated with EGF. • EGF-induced activation of reserve cells is dependent on sphingosine kinase and ERK. • The S1P receptor S1P2 is involved in EGF-induced reserve cell activation. • EGF-induced reserve cell activation is mediated by S1P and its

  8. Mechanisms of vascular endothelial growth factor-induced pathfinding by endothelial sprouts in biomaterials.

    PubMed

    Shamloo, Amir; Xu, Hui; Heilshorn, Sarah

    2012-02-01

    A critical property of biomaterials for use in regenerative medicine applications is the ability to promote angiogenesis, the formation of new vascular networks, to support regenerating tissues. Recent studies have demonstrated that a complex interplay exists between biomechanical and biochemical regulators of endothelial cell sprouting, an early step in angiogenesis. Here, we use a microfluidic platform to study the pathfinding behaviors induced by various stable vascular endothelial growth factor (VEGF) gradients during sprouting morphogenesis within biomaterials. Quantitative, time-lapse analysis of endothelial sprouting demonstrated that the ability of VEGF to regulate sprout orientation during several stages of sprouting morphogenesis (initiation, elongation, and turning navigation) was biomaterial dependent. Identical VEGF gradients induced different types of coordinated cell movements depending on the density of the surrounding collagen/fibronectin matrix. In denser matrices, sprouts were more likely to have an initial orientation aligned parallel to the VEGF gradient. In contrast, in less dense matrices, sprouts were more likely to initially misalign with the VEGF gradient; however, these sprouts underwent significant turning and navigation to eventually reorient to be parallel to the VEGF gradient. These less dense matrices required shallower VEGF gradients and demonstrated lower activating VEGF thresholds to induce proper sprout alignment and pathfinding. These results encourage the future use of microfluidic platforms to probe fundamental aspects of matrix effects on angiogenesis, to screen biomaterials for angiogenic potential, and to design ex vivo tissues with aligned vascular networks.

  9. Mechanisms of Vascular Endothelial Growth Factor-Induced Pathfinding by Endothelial Sprouts in Biomaterials

    PubMed Central

    Shamloo, Amir; Xu, Hui

    2012-01-01

    A critical property of biomaterials for use in regenerative medicine applications is the ability to promote angiogenesis, the formation of new vascular networks, to support regenerating tissues. Recent studies have demonstrated that a complex interplay exists between biomechanical and biochemical regulators of endothelial cell sprouting, an early step in angiogenesis. Here, we use a microfluidic platform to study the pathfinding behaviors induced by various stable vascular endothelial growth factor (VEGF) gradients during sprouting morphogenesis within biomaterials. Quantitative, time-lapse analysis of endothelial sprouting demonstrated that the ability of VEGF to regulate sprout orientation during several stages of sprouting morphogenesis (initiation, elongation, and turning navigation) was biomaterial dependent. Identical VEGF gradients induced different types of coordinated cell movements depending on the density of the surrounding collagen/fibronectin matrix. In denser matrices, sprouts were more likely to have an initial orientation aligned parallel to the VEGF gradient. In contrast, in less dense matrices, sprouts were more likely to initially misalign with the VEGF gradient; however, these sprouts underwent significant turning and navigation to eventually reorient to be parallel to the VEGF gradient. These less dense matrices required shallower VEGF gradients and demonstrated lower activating VEGF thresholds to induce proper sprout alignment and pathfinding. These results encourage the future use of microfluidic platforms to probe fundamental aspects of matrix effects on angiogenesis, to screen biomaterials for angiogenic potential, and to design ex vivo tissues with aligned vascular networks. PMID:21888475

  10. Epidermal growth factor-induced hydrogen peroxide production is mediated by dual oxidase 1.

    PubMed

    Sirokmány, Gábor; Pató, Anna; Zana, Melinda; Donkó, Ágnes; Bíró, Adrienn; Nagy, Péter; Geiszt, Miklós

    2016-08-01

    Stimulation of mammalian cells by epidermal growth factor (EGF) elicits complex signaling events, including an increase in hydrogen peroxide (H2O2) production. Understanding the significance of this response is limited by the fact that the source of EGF-induced H2O2 production is unknown. Here we show that EGF-induced H2O2 production in epidermal cell lines is dependent on the agonist-induced calcium signal. We analyzed the expression of NADPH oxidase isoforms and found both A431 and HaCaT cells to express the calcium-sensitive NADPH oxidase, Dual oxidase 1 (Duox1) and its protein partner Duox activator 1 (DuoxA1). Inhibition of Duox1 expression by small interfering RNAs eliminated EGF-induced H2O2 production in both cell lines. We also demonstrate that H2O2 production by Duox1 leads to the oxidation of thioredoxin-1 and the cytosolic peroxiredoxins. Our observations provide evidence for a new signaling paradigm in which changes of intracellular calcium concentration are transformed into redox signals through the calcium-dependent activation of Duox1. Copyright © 2016. Published by Elsevier Inc.

  11. Flavonoids from the leaves of Carya cathayensis Sarg. inhibit vascular endothelial growth factor-induced angiogenesis.

    PubMed

    Tian, Sha-Sha; Jiang, Fu-Sheng; Zhang, Kun; Zhu, Xue-Xin; Jin, Bo; Lu, Jin-Jian; Ding, Zhi-Shan

    2014-01-01

    The total flavonoids (TFs) were isolated from the leaves of Carya cathayensis Sarg. (LCC), a well-known Chinese medicinal herb commercially cultivated in Tianmu Mountain district, a cross area of Zhejiang and Anhui provinces in China. Five flavonoids, i.e. cardamonin, pinostrobin chalcone (PC), wogonin, chrysin, and pinocembrin were the main components of the TFs. The TFs and these pure compounds suppressed vascular endothelial growth factor (VEGF)-induced angiogenesis as detected in the mouse aortic ring assay, and cardamonin showed the best effect among them. To further elucidate the mechanisms for suppressing angiogenesis of these flavonoids, assays of VEGF-induced proliferation and migration in human umbilical vein endothelial cells (HUVECs) were performed. The TFs, cardamonin, pinocembrin, and chrysin obviously suppressed both VEGF-induced HUVEC proliferation and migration. However, PC and wogonin not only slightly inhibited VEGF-induced proliferation but also remarkably suppressed those of migration in HUVECs. Our further study showed that cardamonin decreased the phosphorylation of ERK and AKT induced by VEGF with a dose-dependent manner in HUVECs. Our findings indicate that the TFs and these pure flavonoids may become potential preventive and/or therapeutic agents against angiogenesis-related diseases.

  12. Abi-1 forms an epidermal growth factor-inducible complex with Cbl: role in receptor endocytosis.

    PubMed

    Tanos, Barbara E; Pendergast, Ann Marie

    2007-07-01

    The Abl-interactor (Abi) proteins are involved in the regulation of actin polymerization and have recently been shown to modulate epidermal growth factor receptor (EGFR) endocytosis. Here we describe the identification of a novel complex between Abi-1 and the Cbl ubiquitin ligase that is induced by stimulation with EGF. Notably, an Abi-1 mutant lacking the SH3 domain (DeltaSH3) fails to interact with Cbl and inhibits EGFR internalization. We show that expression of the Abi-1DeltaSH3 mutant inhibits Cbl accumulation at the plasma membrane after EGF treatment. We have previously shown that the oncogenic Abl tyrosine kinase inhibits EGFR internalization. Here we report that the oncogenic Abl kinase disrupts the EGF-inducible Abi-1/Cbl complex, highlighting the importance of Abl kinases and downstream effectors in the regulation of EGFR internalization. Thus, our work reveals a new role for oncogenic Abl tyrosine kinases in the regulation of the Abi-1/Cbl protein complex and uncovers a role for the Abi-1/Cbl complex in the regulation of EGFR endocytosis.

  13. PLEIOTROPHIN, A MULTIFUNCTIONAL CYTOKINE AND GROWTH FACTOR, INDUCES LEUKOCYTE RESPONSES THROUGH THE INTEGRIN MAC-1.

    PubMed

    Shen, Di; Podolnikova, Nataly P; Yakubenko, Valentin P; Ardell, Christopher L; Balabiyev, Arnat; Ugarova, Tatiana P; Wang, Xu

    2017-09-22

    Pleiotrophin (PTN) is a multifunctional, cationic, glycosaminoglycan-binding cytokine and growth factor involved in numerous physiological and pathological processes, including tissue repair and inflammation-related diseases. PTN has been shown to promote leukocyte responses by inducing their migration and expression of inflammatory cytokines. However, the mechanisms through which PTN mediates these responses remain unclear. Here, we identified the integrin Mac-1 (αMβ2, CD11b/CD18) as the receptor mediating macrophage adhesion and migration to PTN. We also found that expression of Mac-1 on the surface of human embryonic kidney (HEK) 293 cells induced their adhesion and migration to PTN. Accordingly, PTN promoted Mac-1-dependent cell spreading and initiated intracellular signaling manifested in phosphorylation of Erk1/2. While binding to PTN, Mac-1 on Mac-1-expressing HEK293 cells appear to cooperate with cell-surface proteoglycans, since both anti-Mac-1 function-blocking mAb and heparin were required to block adhesion. Moreover, biolayer interferometry and NMR indicated a direct interaction between the αMI domain, the major ligand-binding region of Mac-1, and PTN. Using peptide libraries, we found that in PTN, the αMI domain bound sequences enriched in basic and hydrophobic residues, indicating that PTN conforms to the general principle of ligand-recognition specificity of the αMI domain toward cationic proteins/peptides. Finally, using recombinant PTN-derived fragments, we show that PTN contains two distinct Mac-1-binding sites in each of its constitutive domains. Collectively, these results identify PTN as a ligand for the integrin Mac-1 on the surface of leukocytes and suggest that this interaction may play a role in inflammatory responses. Copyright © 2017, The American Society for Biochemistry and Molecular Biology.

  14. Sphingosine kinase activation regulates hepatocyte growth factor induced migration of endothelial cells.

    PubMed

    Duan, Hai-Feng; Wu, Chu-Tse; Lu, Ying; Wang, Hua; Liu, Hong-Jun; Zhang, Qun-Wei; Jia, Xiang-Xu; Lu, Zhu-Zhuang; Wang, Li-Sheng

    2004-08-15

    Hepatocyte growth factor (HGF)-induced migration of endothelial cells is critical for angiogenesis. Sphingosine kinase (SPK) is a key enzyme catalyzing the formation of sphingosine-1-phosphate (S1P), a lipid messenger that is implicated in the regulation of a wide variety of important cellular events through both intracellular and extracellular mechanisms. The aim of this study was to investigate whether activation of SPK is involved in the migration of endothelial cells induced by HGF. The biological functions of HGF are mediated through the activation of its high-affinity tyrosine kinase receptor, c-met protooncogene. In the present study, Treatment of ECV304 endothelial cells with HGF resulted in tyrosine phosphorylation of c-Met and activation of SPK in a concentration-dependent manner. Either Ly294002 or PD98059, specific inhibitor of the PI3K and ERK/MAPK pathways, respectively, blocked the HGF-induced activation of SPK. HGF stimulation significantly increased intracellular S1P level, but no detectable secretion of S1P into the cell culture medium was observed. Treatment of ECV304 cells with pertussis toxin (PTX) has no effect on the HGF-induced migration, indicating extracellular S1P is dispensable for this process. Overexpression of wild-type SPK gene in ECV 304 cells increased the intracellular S1P and enhanced the HGF-induced migration, whereas inhibition of cellular SPK activity by N,N-dimethylsphingosine (DMS), a potent inhibitor of SPK, or by expression of a dominant-negative SPK (DN-SK) blocked the HGF-induced migration of ECV 304 cells. It is suggested that PI3K and ERK/MAPK mediated the activation of SPK and would be involved in the HGF-induced migration of endothelial cells. These results elucidate a novel mechanism by which intracellularly generated S1P mediates signaling from HGF/c-Met to the endothelial cell migration.

  15. Trichomonas vaginalis homolog of macrophage migration inhibitory factor induces prostate cell growth, invasiveness, and inflammatory responses.

    PubMed

    Twu, Olivia; Dessí, Daniele; Vu, Anh; Mercer, Frances; Stevens, Grant C; de Miguel, Natalia; Rappelli, Paola; Cocco, Anna Rita; Clubb, Robert T; Fiori, Pier Luigi; Johnson, Patricia J

    2014-06-03

    The human-infective parasite Trichomonas vaginalis causes the most prevalent nonviral sexually transmitted infection worldwide. Infections in men may result in colonization of the prostate and are correlated with increased risk of aggressive prostate cancer. We have found that T. vaginalis secretes a protein, T. vaginalis macrophage migration inhibitory factor (TvMIF), that is 47% similar to human macrophage migration inhibitory factor (HuMIF), a proinflammatory cytokine. Because HuMIF is reported to be elevated in prostate cancer and inflammation plays an important role in the initiation and progression of cancers, we have explored a role for TvMIF in prostate cancer. Here, we show that TvMIF has tautomerase activity, inhibits macrophage migration, and is proinflammatory. We also demonstrate that TvMIF binds the human CD74 MIF receptor with high affinity, comparable to that of HuMIF, which triggers activation of ERK, Akt, and Bcl-2-associated death promoter phosphorylation at a physiologically relevant concentration (1 ng/mL, 80 pM). TvMIF increases the in vitro growth and invasion through Matrigel of benign and prostate cancer cells. Sera from patients infected with T. vaginalis are reactive to TvMIF, especially in males. The presence of anti-TvMIF antibodies indicates that TvMIF is released by the parasite and elicits host immune responses during infection. Together, these data indicate that chronic T. vaginalis infections may result in TvMIF-driven inflammation and cell proliferation, thus triggering pathways that contribute to the promotion and progression of prostate cancer.

  16. Neuroprotective effects of ultrasound-guided nerve growth factor injections after sciatic nerve injury

    PubMed Central

    Li, Hong-fei; Wang, Yi-ru; Huo, Hui-ping; Wang, Yue-xiang; Tang, Jie

    2015-01-01

    Nerve growth factor (NGF) plays an important role in promoting neuroregeneration after peripheral nerve injury. However, its effects are limited by its short half-life; it is therefore important to identify an effective mode of administration. High-frequency ultrasound (HFU) is increasingly used in the clinic for high-resolution visualization of tissues, and has been proposed as a method for identifying and evaluating peripheral nerve damage after injury. In addition, HFU is widely used for guiding needle placement when administering drugs to a specific site. We hypothesized that HFU guiding would optimize the neuroprotective effects of NGF on sciatic nerve injury in the rabbit. We performed behavioral, ultrasound, electrophysiological, histological, and immunohistochemical evaluation of HFU-guided NGF injections administered immediately after injury, or 14 days later, and compared this mode of administration with intramuscular NGF injections. Across all assessments, HFU-guided NGF injections gave consistently better outcomes than intramuscular NGF injections administered immediately or 14 days after injury, with immediate treatment also yielding better structural and functional results than when the treatment was delayed by 14 days. Our findings indicate that NGF should be administered as early as possible after peripheral nerve injury, and highlight the striking neuroprotective effects of HFU-guided NGF injections on peripheral nerve injury compared with intramuscular administration. PMID:26807123

  17. Neuroprotective effects of ultrasound-guided nerve growth factor injections after sciatic nerve injury.

    PubMed

    Li, Hong-Fei; Wang, Yi-Ru; Huo, Hui-Ping; Wang, Yue-Xiang; Tang, Jie

    2015-11-01

    Nerve growth factor (NGF) plays an important role in promoting neuroregeneration after peripheral nerve injury. However, its effects are limited by its short half-life; it is therefore important to identify an effective mode of administration. High-frequency ultrasound (HFU) is increasingly used in the clinic for high-resolution visualization of tissues, and has been proposed as a method for identifying and evaluating peripheral nerve damage after injury. In addition, HFU is widely used for guiding needle placement when administering drugs to a specific site. We hypothesized that HFU guiding would optimize the neuroprotective effects of NGF on sciatic nerve injury in the rabbit. We performed behavioral, ultrasound, electrophysiological, histological, and immunohistochemical evaluation of HFU-guided NGF injections administered immediately after injury, or 14 days later, and compared this mode of administration with intramuscular NGF injections. Across all assessments, HFU-guided NGF injections gave consistently better outcomes than intramuscular NGF injections administered immediately or 14 days after injury, with immediate treatment also yielding better structural and functional results than when the treatment was delayed by 14 days. Our findings indicate that NGF should be administered as early as possible after peripheral nerve injury, and highlight the striking neuroprotective effects of HFU-guided NGF injections on peripheral nerve injury compared with intramuscular administration.

  18. Nerve Growth Factor Decreases in Sympathetic and Sensory Nerves of Rats with Chronic Heart Failure

    PubMed Central

    Lu, Jian

    2014-01-01

    Nerve growth factor (NGF) plays a critical role in the maintenance and survival of both sympathetic and sensory nerves. Also, NGF can regulate receptor expression and neuronal activity in the sympathetic and sensory neurons. Abnormalities in NGF regulation are observed in patients and animals with heart failure (HF). Nevertheless, the effects of chronic HF on the levels of NGF within the sympathetic and sensory nerves are not known. Thus, the ELISA method was used to assess the levels of NGF in the stellate ganglion (SG) and dorsal root ganglion (DRG) neurons of control rats and rats with chronic HF induced by myocardial infarction. Our data show for the first time that the levels of NGF were significantly decreased (P < 0.05) in the SG and DRG neurons 6–20 weeks after ligation of the coronary artery. In addition, a close relation was observed between the NGF levels and the left ventricular function. In conclusion, chronic HF impairs the expression of NGF in the sympathetic and sensory nerves. Given that sensory afferent nerves are engaged in the sympathetic nervous responses to somatic stimulation (i.e. muscle activity during exercise) via a reflex mechanism, our data indicate that NGF is likely responsible for the development of muscle reflex-mediated abnormal sympathetic responsiveness observed in chronic HF. PMID:24913185

  19. Nerve Growth Factor: A Focus on Neuroscience and Therapy

    PubMed Central

    Aloe, Luigi; Rocco, Maria Luisa; Omar Balzamino, Bijorn; Micera, Alessandra

    2015-01-01

    Nerve growth factor (NGF) is the firstly discovered and best characterized neurotrophic factor, known to play a critical protective role in the development and survival of sympathetic, sensory and forebrain cholinergic neurons. NGF promotes neuritis outgrowth both in vivo and in vitro and nerve cell recovery after ischemic, surgical or chemical injuries. Recently, the therapeutic property of NGF has been demonstrated on human cutaneous and corneal ulcers, pressure ulcer, glaucoma, maculopathy and retinitis pigmentosa. NGF eye drops administration is well tolerated, with no detectable clinical evidence of systemic or local adverse effects. The aim of this review is to summarize these biological properties and the potential clinical development of NGF. PMID:26411962

  20. Nerve growth factor gene therapy in Alzheimer disease.

    PubMed

    Tuszynski, Mark H

    2007-01-01

    Nervous system growth factors potently stimulate cell function and prevent neuronal death. These broad effects on survival and function arise from direct downstream activation of antiapoptotic pathways, inhibition of proapoptotic pathways, and stimulation of functionally important cellular mechanisms including ERK/MAP kinase and CREB. Thus, as a class, growth factors offer the potential to treat neurodegenerative disorders for the first time by preventing neuronal degeneration rather than compensating for cell loss after it has occurred. Different growth factors affect distinct and specific populations of neurons: the first nervous system growth factor identified, nerve growth factor, potentially stimulates the survival and function of basal forebrain cholinergic neurons, suggesting that nerve growth factor could be a means for reducing the cholinergic component of cell degeneration in Alzheimer disease. This review will discuss the transition of growth factors from preclinical studies to human clinical trials in Alzheimer disease. The implementation of clinical testing of growth factor therapy for neurologic disease has been constrained by the dual need to achieve adequate concentrations of these proteins in specific brain regions containing degenerating neurons, and preventing growth factor spread to nontargeted regions to avoid adverse effects. Gene therapy is one of a limited number of potential methods for achieving these requirements.

  1. Biochemical and biological properties of the nerve growth factor receptor

    SciTech Connect

    Taniuchi, M.

    1988-01-01

    We have utilized a monoclonal antibody (192-IgG) to study the rat nerve growth factor receptor. After intraocular injection, {sup 125}I-192-IgG was retrogradely transported in sympathetic neuronal axons to the superior cervical ganglion. When the sciatic nerve was ligated to induce the accumulation of axonally transported materials, 192-IgG immunostaining was observed on both sides of the ligature, indicating that NGF receptors are transported in both orthograde and retrograde directions. By using {sup 125}I-NGF crosslinking and 192-IgG immunoprecipitation, we detected receptor molecules throughout the rat brain, thereby supporting the hypothesis that NGF is active in the central nervous system. We also discovered that sciatic nerve transection leads to a dramatic increase in the amount of NGF receptor found in the distal portion of the nerve. Immunostaining revealed that all Schwann cells in the distal axotomized nerve were expressing NGF receptors. We examined phosphorylation of NGF receptor in cultured sympathetic neurons and PC12 cells. We also examined pharmacological effects of 192-IgG. Systemic injection of 192-IgG into neonatal rats caused a permanent partial sympathectomy in a dose-dependent manner; a maximum of 50% of the cells were killed.

  2. Heparin Inhibits Hepatocyte Growth Factor Induced Motility and Invasion of Hepatocellular Carcinoma Cells through Early Growth Response Protein 1

    PubMed Central

    Ozen, Evin; Gozukizil, Aysim; Erdal, Esra; Uren, Aykut; Bottaro, Donald P.; Atabey, Nese

    2012-01-01

    The Hepatocyte Growth Factor (HGF)/c-Met signaling pathway regulates hepatocyte proliferation, and pathway aberrations are implicated in the invasive and metastatic behaviors of hepatocellular carcinoma (HCC). In addition to c-Met, heparin acts as a co-receptor to modulate pathway activity. Recently, anti-metastatic and anti-cancer effects of heparin have been reported. However, the role of heparin in the regulation of HGF signaling remains controversial and the effects of heparin on HGF-induced biological responses during hepatocarcinogenesis is not yet defined. In this study we determined the effects of heparin on HGF-induced activities of HCC cells and the underlying molecular mechanisms. Here, we report for the first time that heparin inhibits HGF-induced adhesion, motility and invasion of HCC cells. In addition, heparin reduced HGF-induced activation of c-Met and MAPK in a dose-dependent manner, as well as decreased transcriptional activation and expression of Early growth response factor 1 (Egr1). HGF-induced MMP-2 and MMP-9 activation, and MT1-MMP expression, also were inhibited by heparin. Stable knockdown of Egr1 caused a significant decrease in HGF-induced invasion, as well as the activation and expression of MMPs. Parallel to these findings, the overexpression of Egr1 increased the invasiveness of HCC cells. Our results suggest that Egr1 activates HGF-induced cell invasion through the regulation of MMPs in HCC cells and heparin inhibits HGF-induced cellular invasion via the downregulation of Egr1. Therefore, heparin treatment might be a therapeutic approach to inhibit invasion and metastasis of HCC, especially for patients with active HGF/c-Met signaling. PMID:22912725

  3. Polyethlyene glycol microgels to deliver bioactive nerve growth factor.

    PubMed

    Stukel, Jessica; Thompson, Susan; Simon, Laurent; Willits, Rebecca

    2015-02-01

    Delivery of bioactive molecules is a critical step in fabricating materials for regenerative medicine, yet, this step is particularly challenging in hydrated scaffolds such as hydrogels. Although bulk photocrosslinked poly(ethylene glycol) (PEG) hydrogels have been used for a variety of tissue engineering applications, their capability as drug delivery scaffolds has been limited due to undesirable release profiles and reduction in bioactivity of molecules. To solve these problems, this article presents the fabrication of degradable PEG microgels, which are micron-sized spherical hydrogels, to deliver bioactive nerve growth factor (NGF). NGF release and activity was measured after encapsulation in microgels formed from either 3 kDa or 6 kDa PEG to determine the role of hydrogel mesh size on release. Microgels formed from 6 kDa PEG were statistically larger and had a higher swelling ratio than 3 kDa PEG. The 6 kDa PEG microgels provided a Fickian release with a reduced burst effect and 3 kDa microgels provided anomalous release over ≥20 days. Regardless of molecular weight of PEG, NGF bioactivity was not significantly reduced compared to unprocessed NGF. These results demonstrate that microgels provide easy mechanisms to control the release while retaining the activity of growth factors. As this microgel-based delivery system can be injected at the site of nerve injury to promote nerve repair, the potential to deliver active growth factors in a controlled manner may reduce healing time for neural tissue engineering applications.

  4. Comparison of rabbit facial nerve regeneration in nerve growth factor-containing silicone tubes to that in autologous neural grafts.

    PubMed

    Spector, J G; Lee, P; Derby, A; Roufa, D G

    1995-11-01

    Previous reports suggest that nerve growth factor (NGF) enhanced nerve regeneration in rabbit facial nerves. We compared rabbit facial nerve regeneration in 10-mm silicone tubes prefilled with NGF or cytochrome C (Cyt C), bridging an 8-mm nerve gap, to regeneration of 8-mm autologous nerve grafts. Three weeks following implantation, NGF-treated regenerates exhibited a more mature fascicular organization and more extensive neovascularization than Cyt C-treated controls. Morphometric analysis at the middle of the tube of 3- and 5-week regenerates revealed no significant difference in the mean number of myelinated or unmyelinated axons between NGF- and Cyt C-treated implants. However, when the numbers of myelinated fibers in 5-week regenerates were compared to those in their respective preoperative controls, NGF-treated regenerates had recovered a significantly greater percentage of myelinated axons than Cyt C-treated implants (46% versus 18%, respectively). The number of regenerating myelinated axons in the autologous nerve grafts at 5 weeks was significantly greater than the number of myelinated axons in the silicone tubes. However, in the nerve grafts the majority of the axons were found in the extrafascicular connective tissue (66%). The majority of these myelinated fibers did not find their way into the distal nerve stump. Thus, although the number of regenerating myelinated axons within the nerve grafts is greater than that of axons within silicone tube implants, functional recovery of autologous nerve graft repairs may not be superior to that of intubational repairs.

  5. NERVE GROWTH FACTOR MAINTAINS POTASSIUM CONDUCTANCE AFTER NERVE INJURY IN ADULT CUTANEOUS AFFERENT DORSAL ROOT GANGLION NEURONS

    PubMed Central

    EVERILL, B.; KOCSIS, J. D.

    2008-01-01

    Whole-cell patch-clamp techniques were used to study the effects of nerve growth factor on voltage-dependent potassium conductance in normal and axotomized identified large cutaneous afferent dorsal root ganglion neurons (48–50 μm diameter) many of which probably give rise to myelinated Aβ fibers. K-currents were isolated by blocking Na- and Ca-currents with appropriate ion replacement and channel blockers. Separation of current components was achieved on the basis of response to variation in conditioning voltage. Cutaneous afferents were labeled by the retrograde marker hydroxy-stilbamide (FluoroGold) which was injected into the skin of the foot. The sciatic nerve was either ligated or crushed with fine forceps five to seven days later. Neurons were dissociated 14–17 days after injury. The cut ends of the sciatic nerves were positioned into polyethylene tubes, which were connected to mini-osmotic pumps filled with either nerve growth factor or sterile saline. Control neurons displayed a prominent sustained K-current and the transient potassium currents “A” and “D”. Nerve ligation, which blocks target reconnection resulted in near 50% reduction of total outward current; isolated sustained K-current and transient A-current were reduced by a comparable amount. Nerve crush, which allows regeneration to peripheral targets and exposure of the regenerating nerve to the distal nerve segment, resulted in a small reduction in sustained K-current but no reduction in transient A-current compared to controls. Levels of transient A-current and sustained K-current were maintained at control levels after nerve growth factor treatment. These results indicate that the large reduction in transient A-current, and in sustained K-current, observed in cutaneous afferent cell bodies after nerve ligation is prevented by application of nerve growth factor. PMID:11008179

  6. Sequential activation of phosphatidylinositol 3-kinase, beta Pix, Rac1, and Nox1 in growth factor-induced production of H2O2.

    PubMed

    Park, Hye Sun; Lee, Seung Hye; Park, Dongeun; Lee, Jun Sung; Ryu, Sung Ho; Lee, Won Jae; Rhee, Sue Goo; Bae, Yun Soo

    2004-05-01

    The generation of reactive oxygen species (ROS) in cells stimulated with growth factors requires the activation of phosphatidylinositol 3-kinase (PI3K) and the Rac protein. We report here that the COOH-terminal region of Nox1, a protein related to gp91(phox) (Nox2) of phagocytic cells, is constitutively associated with beta Pix, a guanine nucleotide exchange factor for Rac. Both growth factor-induced ROS production and Rac1 activation were completely blocked in cells depleted of beta Pix by RNA interference. Rac1 was also shown to bind to the COOH-terminal region of Nox1 in a growth factor-dependent manner. Moreover, the depletion of Nox1 by RNA interference inhibited growth factor-induced ROS generation. These results suggest that ROS production in growth factor-stimulated cells is mediated by the sequential activation of PI3K, beta Pix, and Rac1, which then binds to Nox1 to stimulate its NADPH oxidase activity.

  7. Discovery of SNPs in the swine nerve growth factor gene.

    PubMed

    Chung, H Y; Kim, J Y

    2010-10-01

    This study was aimed to search genetic variants for the swine nerve growth factor gene that associated with regulation of proliferation and differentiation of nervous systems. The swine nerve growth factor gene was screened with 5 primer sets for random populations of crossbred pigs born 2005-2007 at National Institute of Animal Science (NIAS). To verify genetic variants of miniature pigs, a total of 288,000 BAC clones generated from NIAS in 2007 were used. The selection of primer sequences was based on sequences of the swine in GenBank (L31898), and genetic variants have been discovered in the crossbred population positioned at 381 (A/C), 412 (C/T), 422 (G/A), 468 (G/C), 496 (A/G), 538 (T/C), 540 (G/A), and 547 (A/G) showing substitutions of amino acids. The identified sequences of miniature pigs including SNPs were submitted into GenBank with an accession number (GQ423508). The sequence alignment conducted to compare genetic distances between species, revealing not many high similarities between swine and human as approximately 0.89 that was a little bit high value than expected. Consequently, we suggest that the identified SNPs of the swine NGF gene may be used in the future to identify genetic markers in coding regions, regarding explanations of phenotypic variations.

  8. Neurotrophic Factor Receptor Expression and in vitro Nerve Growth of Geniculate Ganglion Neurons That Supply Divergent Nerves

    PubMed Central

    Yamout, Adam; Spec, Andrej; Cosmano, Jason; Kashyap, Manoj; Rochlin, M. William

    2016-01-01

    We investigated which neurotrophic factors may contribute to the divergence of two peripheral nerves emanating from the geniculate ganglion. We compared receptor mRNA profiles of the neurons that supply the nerves, and also the growth of their neurites in response to neurotrophic factors in culture. Three mRNAs, Gfra2, TrkA, and TrkC, were differentially expressed. Only one ligand, Neurturin, promoted substantially different nerve regrowth from the nerves, and therefore may contribute to nerve divergence. Three receptor mRNAs were expressed in 100% of the neurons: TrkB, TrkB.T2 (kinaselacking isoform), and NCAM-140. Ligands for these Trks and FRα-1 promoted more outgrowth than ligands for the other receptors. NT-3 and BDNF synergistically promoted outgrowth. Finally, receptors are coexpressed at random rates, arguing against the existence of neuronal subtypes defined by a combinatorial code of these receptors. PMID:16137986

  9. Induction of nerve growth factor receptors on cultured human melanocytes

    SciTech Connect

    Peacocke, M.; Yaar, M.; Mansur, C.P.; Chao, M.V.; Gilchrest, B.A. )

    1988-07-01

    Normal differentiation and malignant transformation of human melanocytes involve a complex series of interactions during which both genetic and environmental factors play roles. At present, the regulation of these processes is poorly understood. The authors have induced the expression of nerve growth factor (NGF) receptors on cultured human melanocytes with phorbol 12-tetradecanoate 13-acetate and have correlated this event with the appearance of a more differentiated, dendritic morphology. Criteria for NGF receptor expression included protein accumulation and cell-surface immunofluorescent staining with a monoclonal antibody directed against the human receptor and induction of the messenger RNA species as determined by blot-hybridization studies. The presence of the receptor could also be induced by UV irradiation or growth factor deprivation. The NGF receptor is inducible in cultured human melanocytes, and they suggest that NGF may modulate the behavior of this neural crest-derived cell in the skin.

  10. Nerve growth factor combined with an epineural conduit for bridging a short nerve gap (10 mm). A study in rabbits.

    PubMed

    Barmpitsioti, Antonia; Konofaos, Petros; Ignatiadis, Ioannis; Papalois, Apostolos; Zoubos, Aristides B; Soucacos, Panagiotis N

    2011-10-01

    The purpose of this study was to evaluate the effect of direct administration of nerve growth factor (NGF) into an epineural conduit across a short nerve gap (10 mm) in a rabbit sciatic nerve model. The animals were divided into two groups. In group 1, n = 6, a 10-mm defect was created in the sciatic nerve and bridged with an epineural flap. A dose of 1 μg of NGF was locally administered daily for the first 21 days. NGF administration was made inside the epineural flap using a silicone reservoir connected to a silicone tube. In group 2, n = 6, the 10-mm defect was bridged with a nerve graft. This group did not receive any further treatment. At 13 weeks, all animals, before euthanasia, underwent electromyography (EMG) studies and then specimen sent for histology morphometric analysis. NGF administration ensured a significantly increased average number of myelinated axons per μm(2) (P = 0.028) and promoted fiber maturation (P = 0.031) and better EMG results (P = 0.046 for latency P = 0.048 for amplitude), compared with the control group. Although nerve grafts remain the gold standard for peripheral nerve repair, NGF-treated epineural conduits represent a good alternative, particularly when an unfavorable environment for nerve grafts is present.

  11. A Review of Bioactive Release from Nerve Conduits as a Neurotherapeutic Strategy for Neuronal Growth in Peripheral Nerve Injury

    PubMed Central

    Choonara, Yahya E.; Bijukumar, Divya; du Toit, Lisa C.

    2014-01-01

    Peripheral nerve regeneration strategies employ the use of polymeric engineered nerve conduits encompassed with components of a delivery system. This allows for the controlled and sustained release of neurotrophic growth factors for the enhancement of the innate regenerative capacity of the injured nerves. This review article focuses on the delivery of neurotrophic factors (NTFs) and the importance of the parameters that control release kinetics in the delivery of optimal quantities of NTFs for improved therapeutic effect and prevention of dose dumping. Studies utilizing various controlled-release strategies, in attempt to obtain ideal release kinetics, have been reviewed in this paper. Release strategies discussed include affinity-based models, crosslinking techniques, and layer-by-layer technologies. Currently available synthetic hollow nerve conduits, an alternative to the nerve autografts, have proven to be successful in the bridging and regeneration of primarily the short transected nerve gaps in several patient cases. However, current research emphasizes on the development of more advanced nerve conduits able to simulate the effectiveness of the autograft which includes, in particular, the ability to deliver growth factors. PMID:25143934

  12. Peripheral Nerve Repair in Rats Using Composite Hydrogel-Filled Aligned Nanofiber Conduits with Incorporated Nerve Growth Factor

    PubMed Central

    Jin, Jenny; Limburg, Sonja; Joshi, Sunil K.; Landman, Rebeccah; Park, Michelle; Zhang, Qia; Kim, Hubert T.

    2013-01-01

    Repair of peripheral nerve defects with current synthetic, tubular nerve conduits generally shows inferior recovery when compared with using nerve autografts, the current gold standard. We tested the ability of composite collagen and hyaluronan hydrogels, with and without the nerve growth factor (NGF), to stimulate neurite extension on a promising aligned, nanofiber poly-L-lactide-co-caprolactone (PLCL) scaffold. In vitro, the hydrogels significantly increased neurite extension from dorsal root ganglia explants. Consistent with these results, the addition of hydrogels as luminal fillers within aligned, nanofiber tubular PLCL conduits led to improved sensory function compared to autograft repair in a critical-size defect in the sciatic nerve in a rat model. Sensory recovery was assessed 3 and 12 weeks after repair using a withdrawal assay from thermal stimulation. The addition of hydrogel did not enhance recovery of motor function in the rat model. The NGF led to dose-dependent improvements in neurite out-growth in vitro, but did not have a significant effect in vivo. In summary, composite collagen/hyaluronan hydrogels enhanced sensory neurite outgrowth in vitro and sensory recovery in vivo. The use of such hydrogels as luminal fillers for tubular nerve conduits may therefore be useful in assisting restoration of protective sensation following peripheral nerve injury. PMID:23659607

  13. Efficacy of glial growth factor and nerve growth factor on the recovery of traumatic facial paralysis.

    PubMed

    Yildiz, Mucahit; Karlidag, Turgut; Yalcin, Sinasi; Ozogul, Candan; Keles, Erol; Alpay, Hayrettin Cengiz; Yanilmaz, Muhammed

    2011-08-01

    The aim of this study was to assess the effects of Glial growth factor (GGF) and nerve growth factor (NGF) on nerve regeneration in facial nerve anastomosis. In this study, approximately a 1-mm segment was resected from the facial nerve and the free ends were anastomosed. All animals underwent the same surgical procedure and 30 rabbits were grouped randomly in three groups. Control group, the group without any medications; NGF group, the group receiving 250 ng/0.1 ml NGF in the epineurium at the site of anastomosis; GBF group, the group receiving 500 ng/0.1 ml GGF in the epineurium at the site of anastomosis. Medications were given at the time of surgery, and at 24 and 48 h postoperatively. After 2 months, the sites of anastomosis were excised and examined using the electron microscope. It was found that the best regeneration was in the group receiving GGF as compared to the control group in terms of nerve regeneration. Schwann cell and glial cell proliferation were found to be significantly higher in the group receiving GGF as compared to the group receiving NGF. Besides, the number of myelin debris, an indicator of degeneration, was significantly lower in the group with GGF as compared to NGF and control groups (p < 0.005). Using GGF and NGF in order to increase regeneration after nerve anastomosis in experimental traumatic facial nerve paralysis may be a hopeful alternative treatment option in the future. However, further studies on human studies are required to support these results.

  14. Mast Cells Synthesize, Store, and Release Nerve Growth Factor

    NASA Astrophysics Data System (ADS)

    Leon, A.; Buriani, A.; dal Toso, R.; Fabris, M.; Romanello, S.; Aloe, L.; Levi-Montalcini, R.

    1994-04-01

    Mast cells and nerve growth factor (NGF) have both been reported to be involved in neuroimmune interactions and tissue inflammation. In many peripheral tissues, mast cells interact with the innervating fibers. Changes in the behaviors of both of these elements occur after tissue injury/inflammation. As such conditions are typically associated with rapid mast cell activation and NGF accumulation in inflammatory exudates, we hypothesized that mast cells may be capable of producing NGF. Here we report that (i) NGF mRNA is expressed in adult rat peritoneal mast cells; (ii) anti-NGF antibodies clearly stain vesicular compartments of purified mast cells and mast cells in histological sections of adult rodent mesenchymal tissues; and (iii) medium conditioned by peritoneal mast cells contains biologically active NGF. Mast cells thus represent a newly recognized source of NGF. The known actions of NGF on peripheral nerve fibers and immune cells suggest that mast cell-derived NGF may control adaptive/reactive responses of the nervous and immune systems toward noxious tissue perturbations. Conversely, alterations in normal mast cell behaviors may provoke maladaptive neuroimmune tissue responses whose consequences could have profound implications in inflammatory disease states, including those of an autoimmune nature.

  15. Topical administration of adrenergic receptor pharmaceutics and nerve growth factor

    PubMed Central

    Steinle, Jena J

    2010-01-01

    Topical application of nerve growth factor (NGF) and adrenergic receptor pharmaceutics are currently in use for corneal ulcers and glaucoma. A recent interest in the neuroprotective abilities of NGF has led to a renewed interest in NGF as a therapeutic for retinal and choroidal diseases. NGF can promote cell proliferation through actions of the TrkA receptor or promote apoptosis through receptor p75NTR. This understanding has led to novel interest in the role of NGF for diseases of the posterior eye. The role of β-adrenergic receptor agonists and antagonists for treatments of glaucoma, diabetic retinopathy, and their potential mechanisms of action, are still under investigation. This review discusses the current knowledge and applications of topical NGF and adrenergic receptor drugs for ocular disease. PMID:20668722

  16. Proteasome Inhibition by Fellutamide B Induces Nerve Growth Factor Synthesis

    PubMed Central

    Hines, John; Groll, Michael; Fahnestock, Margaret; Crews, Craig M.

    2008-01-01

    SUMMARY Neurotrophic small molecules have the potential to aid in the treatment of neuronal injury and neurodegenerative diseases. The natural product fellutamide B, originally isolated from Penicillium fellutanum, potently induces nerve growth factor (NGF) release from fibroblasts and glial-derived cells, although the mechanism for this neurotrophic activity has not been elucidated. Here, we report that fellutamide B potently inhibits proteasome catalytic activity. High resolution structural information obtained from co-crystallization of the 20S proteasome reveals novel aspects regarding β-subunit binding and adduct formation by fellutamide B to inhibit their hydrolytic activity. We demonstrate that fellutamide B and other proteasome inhibitors increased NGF gene transcription via a cis-acting element (or elements) in the promoter. These results demonstrate an unrecognized connection between proteasome inhibition and NGF production, suggesting a possible new strategy in the development of neurotrophic agents. PMID:18482702

  17. Nerve growth factor: stimulation of polymorphonuclear leukocyte chemotaxis in vitro.

    PubMed Central

    Gee, A P; Boyle, M D; Munger, K L; Lawman, M J; Young, M

    1983-01-01

    Topical application of mouse nerve growth factor (NGF) to superficial skin wounds of mice has previously been shown to accelerate the rate of wound contraction. Results of the present study reveal that NGF in the presence of plasma is also chemotactic for human polymorphonuclear leukocytes in vitro, and the concentration of NGF required for this effect is similar to that which stimulates ganglionic neurite outgrowth. This property does not arise from liberation of the C5a fragment of complement, nor does it require the known enzymic activity of NGF. (NGF inactivated with diisopropyl fluorophosphate is equally active.) We conclude that NGF can display biological effects on cells of nonneural origin and function, and this feature might play a role in the early inflammatory response to injury. PMID:6580641

  18. Nerve Growth Factor Potentiates the Neurotoxicity of β Amyloid

    NASA Astrophysics Data System (ADS)

    Yankner, Bruce A.; Caceres, Alfredo; Duffy, Lawrence K.

    1990-11-01

    The role of growth factors in the pathogenesis of Alzheimer disease is unknown. The β-amyloid protein accumulates abnormally in the brain in Alzheimer disease and is neurotoxic to differentiated hippocampal neurons in culture. Nerve growth factor (NGF) increased the neurotoxic potency of a β-amyloid polypeptide by a factor of ≈100,000, which resulted in a reduction of the β-amyloid neurotoxic EC50 from 0.1 μM to 1 pM. This potentiating effect of NGF was reversed by a monoclonal antibody against NGF and was not observed for a variety of other neurotrophic growth factors. Exposure of hippocampal neurons to very low concentrations of β amyloid alone resulted in a marked induction of immunoreactive NGF receptors. Addition of NGF with β amyloid resulted in the appearance of neurodegenerative changes in NGF receptor-positive neurons. The early and profound degeneration of hippocampal and basal forebrain cholinergic neurons that occurs in Alzheimer disease may result from a neurotoxic interaction of β amyloid with NGF.

  19. [Nerve growth factor in neurodegeneration and neurorestorative therapy].

    PubMed

    Lorigados-Pedre, L; Bergado-Rosado, J

    The purpose of this work was to gather the information currently available about the content of nerve growth factor (NGF) in experimental models of neurodegeneration and in neurodegenerative diseases, including Alzheimer's, Parkinson's and Huntington's diseases, as well as to analyse how NGF content is affected by the application of different neurorestorative therapies (transplant and trophic therapy) in these neurological entities. Neurotrophins are proteins that promote the differentiation, growth and survival of many populations of peripheral neurons and the central nervous system during development and adulthood. NGF is the best known and most widely researched member of this family, which is also made up of the growth factor derived from the brain and neurotrophins 3, 4/5, 6 and 7. In the last few decades, significant progress has been made in the knowledge available about the biological role played by these factors, their molecular characterisation and regulation, as well as their signalling mechanisms. Yet, little is known about the role played by the neurotrophic factors in neurodegenerative diseases or whether the levels of these factors are modified following the use of neurorestorative treatment. Neurodegenerative disorders, especially Parkinson and Alzheimer, are accompanied by modifications in the levels of NGF that depend on the extent to which the disease has progressed. A model of the changes in NGF content during neurodegenerative processes is also proposed.

  20. Intrathecal administration of nerve growth factor delays GAP 43 expression and early phase regeneration of adult rat peripheral nerve.

    PubMed

    Hirata, Akira; Masaki, Toshihiro; Motoyoshi, Kazuo; Kamakura, Keiko

    2002-07-19

    Whether nerve growth factor (NGF) promotes peripheral nerve regeneration in vivo, in particular in adults, is controversial. We therefore examined the effect of exogenous NGF on nerve regeneration and the expression of GAP 43 (growth-associated protein 43) in adult rats. NGF was infused intrathecally via an osmotic mini-pump, while control rats received artificial cerebrospinal fluid. Two days after the infusion was initiated, the right sciatic nerves were transected or crushed, and the animals allowed to survive for 3 to 11 days. The right DRG, the right proximal stump of the transected sciatic nerve, and the posterior horn of the spinal cord were examined by Western blotting, immunohistochemistry, and electron microscopy. GAP 43 immunoreactivity in the NGF-treated animals was significantly lower than in the aCSF-treated controls. Electron microscopy showed that the number of myelinated and unmyelinated axons decreased significantly in the NGF-treated rats as compared with the controls. These findings are indicative that exogenous NGF delayed GAP 43 induction and the early phase of peripheral nerve regeneration and supports the hypothesis that the loss of NGF supply from peripheral targets via retrograde transport caused by axotomy serves as a signal for DRG neurons to invoke regenerative responses. NGF administered intrathecally may delay the neurons' perception of the reduction of the endogenous NGF, causing a delay in conversion of DRG neurons from the normal physiological condition to regrowth state.

  1. alpha-Adrenergic regulation of secretion of mouse saliva rich in nerve growth factor.

    PubMed Central

    Wallace, L J; Partlow, L M

    1976-01-01

    Nerve growth factor has been quantified by both bioassay and radial immunodiffusion in mouse saliva elicited by several secretagogues. The concentrations by bioassay of nerve growth factor in both epinephrine- and norepinephrine-induced saliva (3400 and 900 mug/ml, respectively) are higher than reported in any other source. In contrast, the concentrations of nerve growth factor in isoproterenol- and pilocarpine-induced saliva are relatively low (17 and 2 mug/ml, respectively). The specific activity of the salivary nerve growth factor was 41, 36, 2, and 0.6 mug/mg of protein in secretions elicited by epinephrine, norepinephrine, pilocarpine, and isoproterenol, respectively. Salivation after administration of either epinephrine or norepinephrine was completely inhibited by the alpha-adrenergic blocker, phenoxybenzamine. These results suggest that the release of saliva rich in nerve growth factor is primarily regulated through alpha-adrenergic receptors. Images PMID:186790

  2. Nerve growth factor promotes in vitro proliferation of neural stem cells from tree shrews.

    PubMed

    Xiong, Liu-Lin; Chen, Zhi-Wei; Wang, Ting-Hua

    2016-04-01

    Neural stem cells promote neuronal regeneration and repair of brain tissue after injury, but have limited resources and proliferative ability in vivo. We hypothesized that nerve growth factor would promote in vitro proliferation of neural stem cells derived from the tree shrews, a primate-like mammal that has been proposed as an alternative to primates in biomedical translational research. We cultured neural stem cells from the hippocampus of tree shrews at embryonic day 38, and added nerve growth factor (100 μg/L) to the culture medium. Neural stem cells from the hippocampus of tree shrews cultured without nerve growth factor were used as controls. After 3 days, fluorescence microscopy after DAPI and nestin staining revealed that the number of neurospheres and DAPI/nestin-positive cells was markedly greater in the nerve growth factor-treated cells than in control cells. These findings demonstrate that nerve growth factor promotes the proliferation of neural stem cells derived from tree shrews.

  3. Temporal mismatch between pain behaviour, skin Nerve Growth factor and intra-epidermal nerve fibre density in trigeminal neuropathic pain.

    PubMed

    Evans, Laura J; Loescher, Alison R; Boissonade, Fiona M; Whawell, Simon A; Robinson, Peter P; Andrew, David

    2014-01-01

    The neurotrophin Nerve Growth factor (NGF) is known to influence the phenotype of mature nociceptors, for example by altering synthesis of neuropeptides, and changes in NGF levels have been implicated in the pathophysiology of chronic pain conditions such as neuropathic pain. We have tested the hypothesis that after partial nerve injury, NGF accumulates within the skin and causes 'pro-nociceptive' phenotypic changes in the remaining population of sensory nerve fibres, which could underpin the development of neuropathic pain. Eleven days after chronic constriction injury of the rat mental nerve the intra-epidermal nerve fibre density of the chin skin from had reduced from 11.6 ± 4.9 fibres/mm to 1.0 ± 0.4 fibres/mm; this slowly recovered to 2.4 ± 2.0 fibres/mm on day 14 and 4.0 ± 0.8 fibres/mm on day 21. Cold hyperalgesia in the ipsilateral lower lip was detectable 11 days after chronic constriction injury, although at this time skin [NGF] did not differ between sides. At 14 days post-injury, there was a significantly greater [NGF] ipsilaterally compared to contralaterally (ipsilateral = 111 ± 23 pg/mg, contralateral = 69 ± 13 pg/mg), but there was no behavioural evidence of neuropathic pain at this time-point. By 21 days post-injury, skin [NGF] was elevated bilaterally and there was a significant increase in the proportion of TrkA-positive (the high-affinity NGF receptor) intra-epidermal nerve fibres that were immunolabelled for the neuropeptide Calcitonin Gene-related peptide. The temporal mismatch in behaviour, skin [NGF] and phenotypic changes in sensory nerve fibres indicate that increased [NGF] does not cause hyperalgesia after partial mental nerve injury, although it may contribute to the altered neurochemistry of cutaneous nerve fibres.

  4. Manipulation of the nerve growth factor network in prostate cancer.

    PubMed

    Papatsoris, Athanasios G; Liolitsa, Danae; Deliveliotis, Charalambos

    2007-03-01

    Autocrine and paracrine events regulated by nerve growth factor (NGF) and relevant receptors (low- and high affinity; p75 neurotrophin receptor [p75(NTR)] and TrkA, respectively) seem to play a significant role in prostate carcinogenesis. Studies reveal that p75(NTR) is both a tumor suppressor of growth and a metastasis suppressor of human prostate cancer cells. Furthermore, p75(NTR) is progressively lost during prostate carcinogenesis. An imbalance between p75(NTR) and tropomyosin receptor kinase A (TrkA)-mediated signals may be involved in the progression of prostate cancer through increased proliferation and reduced apoptosis. The antiproliferative and apoptotic effects of GnRH analogs in prostate cancer cells may be mediated by altering the TrkA:p75(NTR) NGF receptor ratio. Administration of NGF induces a reversion of the androgen-independent/androgen receptor-negative prostate cancer cell lines to a less malignant phenotype. Finally, Trk inhibition is a novel, attractive and rational approach for prostate cancer therapy. This review unravels the NGF 'circuitry' in prostate cancinogenesis for relevant pharmacologic manipulation to lead to the development of novel therapeutic agents.

  5. Upregulation of nerve growth factor following cortical trauma.

    PubMed

    DeKosky, S T; Goss, J R; Miller, P D; Styren, S D; Kochanek, P M; Marion, D

    1994-12-01

    As part of the inflammatory response to brain injury, CSF and tissue levels of interleukin-1 beta (IL-1 beta) are elevated after trauma. This elevation in IL-1 beta initiates a cascade of events among which may be an upregulation in nerve growth factor (NGF) in brain tissue. We infused IL-1 beta into the ventricle of adult rats and found a two- to fourfold increase in NGF in the cerebral cortex, hippocampus, and cerebellum, suggesting that IL-1 beta induced in vivo may also increase NGF in the brain. To test this hypothesis we utilized two models of traumatic brain injury (TBI) in the rat and examined NGF protein and RNA in the cortex over a period of several days. Both weight drop and controlled cortical contusion models of CNS trauma demonstrated large and significant increases in NGF protein in the cortex. NGF RNA was assessed in the controlled cortical contusion model and increased approximately fivefold by 1 day post-trauma. The remarkable elevation of NGF observed following TBI suggests that its role in response to injury may be other than as a target-derived growth substance. We hypothesize that the elevation of NGF in trauma induces upregulation of enzymes which suppress free-radical formation after injury.

  6. Nerve growth factor signaling in prostate health and disease.

    PubMed

    Arrighi, Nicola; Bodei, Serena; Zani, Danilo; Simeone, Claudio; Cunico, Sergio Cosciani; Missale, Cristina; Spano, Pierfranco; Sigala, Sandra

    2010-06-01

    The prostate is one of the most abundant sources of nerve growth factor (NGF) in different species, including humans. NGF and its receptors are implicated in the control of prostate cell proliferation and apoptosis and it can either support or suppress cell growth. The co-expression of both NGF receptors, p75(NGFR) and tropomyosin-related kinase A (trkA), represents a crucial condition for the antiproliferative effect of NGF; indeed, p75(NGFR) is progressively lost during prostate tumorigenesis and its disappearance represents a malignancy marker of prostate adenocarcinoma (PCa). Interestingly, a dysregulation of NGF signal transduction was found in a number of human tumors. This review summarizes the current knowledge on the role of NGF and its receptors in prostate and in PCa. Conclusions bring to the hypothesis that the NGF network could be a candidate for future pharmacological manipulation in the PCa therapy: in particular the re-expression of p75(NTR) and/or the negative modulation of trkA could represent a target to induce apoptosis and to reduce proliferation and invasiveness of PCa.

  7. Protein Kinase A Regulates 3-Phosphatidylinositide Dynamics during Platelet-derived Growth Factor-induced Membrane Ruffling and Chemotaxis*S⃞

    PubMed Central

    Deming, Paula B.; Campbell, Shirley L.; Baldor, Linda C.; Howe, Alan K.

    2008-01-01

    Spatial regulation of the cAMP-dependent protein kinase (PKA) is required for chemotaxis in fibroblasts; however, the mechanism(s) by which PKA regulates the cell migration machinery remain largely unknown. Here we report that one function of PKA during platelet-derived growth factor (PDGF)-induced chemotaxis was to promote membrane ruffling by regulating phosphatidylinositol 3,4,5-trisphosphate (PIP3) dynamics. Inhibition of PKA activity dramatically altered membrane dynamics and attenuated formation of peripheral membrane ruffles in response to PDGF. PKA inhibition also significantly decreased the number and size of PIP3-rich membrane ruffles in response to uniform stimulation and to gradients of PDGF. This ruffling defect was quantified using a newly developed method, based on computer vision edge-detection algorithms. PKA inhibition caused a marked attenuation in the bulk accumulation of PIP3 following PDGF stimulation, without effects on PI3-kinase (PI3K) activity. The deficits in PIP3 dynamics correlated with a significant inhibition of growth factor-induced membrane recruitment of endogenous Akt and Rac activation in PKA-inhibited cells. Simultaneous inhibition of PKA and Rac had an additive inhibitory effect on growth factor-induced ruffling dynamics. Conversely, the expression of a constitutively active Rac allele was able to rescue the defect in membrane ruffling and restore the localization of a fluorescent PIP3 marker to membrane ruffles in PKA-inhibited cells, even in the absence of PI3K activity. These data demonstrate that, like Rac, PKA contributes to PIP3 and membrane dynamics independently of direct regulation of PI3K activity and suggest that modulation of PIP3/3-phosphatidylinositol (3-PI) lipids represents a major target for PKA in the regulation of PDGF-induced chemotactic events. PMID:18936099

  8. Recombinant human fibroblast growth factor-2 promotes nerve regeneration and functional recovery after mental nerve crush injury.

    PubMed

    Lee, Sung Ho; Jin, Wei-Peng; Seo, Na Ri; Pang, Kang-Mi; Kim, Bongju; Kim, Soung-Min; Lee, Jong-Ho

    2017-04-01

    Several studies have shown that fibroblast growth factor-2 (FGF2) can directly affect axon regeneration after peripheral nerve damage. In this study, we performed sensory tests and histological analyses to study the effect of recombinant human FGF-2 (rhFGF2) treatment on damaged mental nerves. The mental nerves of 6-week-old male Sprague-Dawley rats were crush-injured for 1 minute and then treated with 10 or 50 μg/mL rhFGF2 or PBS in crush injury area with a mini Osmotic pump. Sensory test using von Frey filaments at 1 week revealed the presence of sensory degeneration based on decreased gap score and increased difference score. However, at 2 weeks, the gap score and difference score were significantly rebounded in the mental nerve crush group treated with 10 μg/mL rhFGF2. Interestingly, treatment with 10 μg/mL rhFGF had a more obviously positive effect on the gap score than treatment with 50 μg/mL rhFGF2. In addition, retrograde neuronal tracing with Dil revealed a significant increase in nerve regeneration in the trigeminal ganglion at 2 and 4 weeks in the rhFGF2 groups (10 μg/mL and 50 μg/mL) than in the PBS group. The 10 μg/mL rhFGF2 group also showed an obviously robust regeneration in axon density in the mental nerve at 4 weeks. Our results demonstrate that 10 μg/mL rhFGF induces mental nerve regeneration and sensory recovery after mental nerve crush injury.

  9. Specific changes in circulating cytokines and growth factors induced by exercise stress testing in asymptomatic aortic valve stenosis.

    PubMed

    Kolasa-Trela, Renata; Konieczynska, Malgorzata; Bazanek, Marta; Undas, Anetta

    2017-01-01

    We evaluated exercise-induced changes in the profile of circulating cytokines and growth factors in patients with AS. We studied 32 consecutive asymptomatic moderate-to-severe AS patients and 32 age and sex-matched controls. Plasma levels of interleukin (IL)-6, IL-10, hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF), and transforming growth factor (TGF)-β were measured at 4 time points, i.e. at rest, at peak bicycle exercise, one hour and 24 hours after a symptom-limited exercise. Exercise increased all the 5 markers in both groups (all p<0.0001). The maximum levels of all tested cytokines were higher in the AS group (all p<0.05) compared with controls. In AS patients the highest levels of VEGF, IL-6, and IL-10 were observed one hour after exercise, while in the control group at peak exercise. In both groups maximum TGF- β levels were observed one hour after exercise. HGF levels were higher at peak and one hour after test in the AS group (p = 0.0001), however the maximum value in AS was observed at peak while in controls after test. In both groups TGF-β was the only marker that remained increased 24 hours after exercise compared with the value at rest (p = 0.0001). The cytokines and growth factors showed no association with heart rate and the workload. In asymptomatic patients with moderate-to-severe AS, exercise produces a different pattern of changes in circulating cytokines and growth factors, and maximum levels of all tested cytokines were significantly higher in AS patients compared with the control group.

  10. Specific changes in circulating cytokines and growth factors induced by exercise stress testing in asymptomatic aortic valve stenosis

    PubMed Central

    Kolasa-Trela, Renata; Konieczynska, Malgorzata; Bazanek, Marta; Undas, Anetta

    2017-01-01

    Background We evaluated exercise-induced changes in the profile of circulating cytokines and growth factors in patients with AS. Methods We studied 32 consecutive asymptomatic moderate-to-severe AS patients and 32 age and sex-matched controls. Plasma levels of interleukin (IL)-6, IL-10, hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF), and transforming growth factor (TGF)-β were measured at 4 time points, i.e. at rest, at peak bicycle exercise, one hour and 24 hours after a symptom-limited exercise. Results Exercise increased all the 5 markers in both groups (all p<0.0001). The maximum levels of all tested cytokines were higher in the AS group (all p<0.05) compared with controls. In AS patients the highest levels of VEGF, IL-6, and IL-10 were observed one hour after exercise, while in the control group at peak exercise. In both groups maximum TGF- β levels were observed one hour after exercise. HGF levels were higher at peak and one hour after test in the AS group (p = 0.0001), however the maximum value in AS was observed at peak while in controls after test. In both groups TGF-β was the only marker that remained increased 24 hours after exercise compared with the value at rest (p = 0.0001). The cytokines and growth factors showed no association with heart rate and the workload. Conclusion In asymptomatic patients with moderate-to-severe AS, exercise produces a different pattern of changes in circulating cytokines and growth factors, and maximum levels of all tested cytokines were significantly higher in AS patients compared with the control group. PMID:28291817

  11. Potential mechanisms for hypoalgesia induced by anti-nerve growth factor immunoglobulin are identified using autoimmune nerve growth factor deprivation

    PubMed Central

    Hoffman, E. Matthew; Zhang, Zijia; Anderson, Michael B.; Schechter, Ruben; Miller, Kenneth E.

    2011-01-01

    Nerve growth factor (NGF) antagonism has long been proposed as a chronic pain treatment. In 2010, the FDA suspended clinical trials using tanezumab, a humanized monoclonal anti-NGF antibody, to treat osteoarthritis due to worsening joint damage in 16 patients. Increased physical activity in the absence of acute pain which normally prevents self harm was purported as a potential cause. Such an adverse effect is consistent with an extension of tanezumab's primary mechanism of action by decreasing pain sensitivity below baseline levels. In animal inflammatory pain models, NGF antagonism decreases intraepidermal nerve fiber (IENF) density and attenuates increases in expression of nociception related proteins, such as calcitonin gene-related peptide (CGRP) and substance P (SP). Little is known of the effects of NGF antagonism in noninflamed animals and the hypoalgesia that ensues. In the current study, we immunized rats with NGF or cytochrome C (cytC) and examined 1) nocifensive behaviors with thermal latencies, mechanical thresholds, the hot plate test, and the tail flick test, 2) IENF density, and 3) expression of CGRP, SP, voltage-gated sodium channel 1.8 (Nav1.8), and glutaminase in subpopulations of dorsal root ganglion (DRG) neurons separated by size and isolectin B4 (IB4) labeling. Rats with high anti-NGF titers had delayed responses on the hot plate test but no other behavioral abnormalities. Delayed hot plate responses correlated with lower IENF density. CGRP and SP expression was decreased principally in medium (400-800 μm2) and small neurons (<400 μm2), respectively, regardless of IB4 labeling. Expression of Nav1.8 was only decreased in small and medium IB4 negative neurons. NGF immunization appears to result in a more profound antagonism of NGF than tanezumab therapy, but we hypothesize that decreases in IENF density and nociception related protein expression are potential mechanisms for tanezumab induced hypoalgesia. PMID:21802499

  12. Regulation by intracellular Ca sup 2+ and cyclic AMP of the growth factor-induced ruffling membrane formation and stimulation of fluid-phase endocytosis and exocytosis

    SciTech Connect

    Miyata, Yoshihiko Tokyo Metropolitan Inst. of Medical Science ); Nishida, Eisuke; Sakai, Hikoichi ); Koyasu, Shigeo; Yahara, Ichiro )

    1989-04-01

    Insulin, insulin-like growth factor-I (IGF-I), and epidermal growth factor (EGF) induce formation of ruffling membranes and stimulate the fluid-phase endocytosis and exocytosis in human epidermoid carcinoma KB cells. An increase in intracellular Ca{sup 2+} concentration by treatment with A23187, a calcium ionophore, or an increase in intracellular cAMP level by treatment with dibutyryl cAMP or forskolin almost completely inhibited the insulin-, IGF-I-, or EGF-induced formation of ruffling membranes. Increases in Ca{sup 2+} or cAMP concentration also inhibited almost completely the stimulation of fluid-phase endocytosis and exocytosis elicited by these growth factors. These results suggest that the growth factor-induced ruffling membrane formation and the stimulation of fluid-phase endocytosis and exocytosis have a common regulatory mechanism involving intracellular concentrations of Ca{sup 2+} and cAMP. {sup 125}I-EGF binding assays and immunoprecipitation experiments with anti-phosphotyrosine antibody revealed that treatment of KB cells with A23187, dibutyryl cAMP, or forskolin did not inhibit the EGF binding to the cells nor subsequent tyrosine autophosphorylation of its receptors. These results indicate that Ca{sup 2+}- and/or cAMP-sensitive intracellular reactions exist downstream from the receptor kinase activation in the process of these early cellular responses.

  13. Anti-nerve growth factor in pain management: current evidence

    PubMed Central

    Chang, David S; Hsu, Eugene; Hottinger, Daniel G; Cohen, Steven P

    2016-01-01

    There continues to be an unmet need for safe and effective pain medications. Opioids and nonsteroidal anti-inflammatory drugs (NSAIDs) dominate the clinical landscape despite limited effectiveness and considerable side-effect profiles. Although significant advancements have identified myriad potential pain targets over the past several decades, the majority of new pain pharmacotherapies have failed to come to market. The discovery of nerve growth factor (NGF) and its interaction with tropomyosin receptor kinase A (trkA) have been well characterized as important mediators of pain initiation and maintenance, and pharmacotherapies targeting this pathway have the potential to be considered promising methods in the treatment of a variety of nociceptive and neuropathic pain conditions. Several methodologic approaches, including sequestration of free NGF, prevention of NGF binding and trkA activation, and inhibition of trkA function, have been investigated in the development of new pharmacotherapies. Among these, NGF-sequestering antibodies have exhibited the most promise in clinical trials. However, in 2010, reports of rapid joint destruction leading to joint replacement prompted the US Food and Drug Administration (FDA) to place a hold on all clinical trials involving anti-NGF antibodies. Although the FDA has since lifted this hold and a number of new trials are under way, the long-term efficacy and safety profile of anti-NGF antibodies are yet to be established. PMID:27354823

  14. Anti-nerve growth factor in pain management: current evidence.

    PubMed

    Chang, David S; Hsu, Eugene; Hottinger, Daniel G; Cohen, Steven P

    2016-01-01

    There continues to be an unmet need for safe and effective pain medications. Opioids and nonsteroidal anti-inflammatory drugs (NSAIDs) dominate the clinical landscape despite limited effectiveness and considerable side-effect profiles. Although significant advancements have identified myriad potential pain targets over the past several decades, the majority of new pain pharmacotherapies have failed to come to market. The discovery of nerve growth factor (NGF) and its interaction with tropomyosin receptor kinase A (trkA) have been well characterized as important mediators of pain initiation and maintenance, and pharmacotherapies targeting this pathway have the potential to be considered promising methods in the treatment of a variety of nociceptive and neuropathic pain conditions. Several methodologic approaches, including sequestration of free NGF, prevention of NGF binding and trkA activation, and inhibition of trkA function, have been investigated in the development of new pharmacotherapies. Among these, NGF-sequestering antibodies have exhibited the most promise in clinical trials. However, in 2010, reports of rapid joint destruction leading to joint replacement prompted the US Food and Drug Administration (FDA) to place a hold on all clinical trials involving anti-NGF antibodies. Although the FDA has since lifted this hold and a number of new trials are under way, the long-term efficacy and safety profile of anti-NGF antibodies are yet to be established.

  15. Nerve growth factor signal transduction in mature pig oligodendrocytes.

    PubMed

    Althaus, H H; Hempel, R; Klöppner, S; Engel, J; Schmidt-Schultz, T; Kruska, L; Heumann, R

    1997-12-01

    It has previously been shown that nerve growth factor (NGF) is of functional significance for mature pig oligodendrocytes (OLs) in culture. The present data give evidence for the expression of TrkA, the so-called high-affinity NGF receptor, and of p75NTR, the so-called low-affinity NGF receptor. TrkA is upregulated during culturing, in contrast to the p75 receptor. Exposure of OLs to NGF induces an autophosphorylation of TrkA via its intrinsic tyrosine kinase. K-252a inhibits the TrkA autophosphorylation, which reduces the OL process formation to control levels. To the tyrosine-phosphorylated sites of TrkA several proteins, such as phospholipase C-gamma1, the adaptor protein SHC, the phosphotyrosine phosphatase SH-PTP2 (SYP) associate via their SH2 phosphotase SH-PTP2 domain. The association of SHC to TrkA is shown by co-immunoprecipitation. Indirect evidence for a possible activation of PLC-gamma1 is given by an NGF-induced increase of oligodendroglial [Ca2+]i. Downstream from TrkA, a mitogen-activated protein kinase cascade, which includes Erk1 and Erk2, is operating. An in-gel myelin basic protein kinase assay revealed that NGF activates predominantly Erk1. Finally, it is shown that NGF stimulates expression of c-fos.

  16. Electroacupucture and nerve growth factor: potential clinical applications.

    PubMed

    Manni, Luigi; Rocco, M L; Barbaro Paparo, Samuele; Guaragna, Morena

    2011-06-01

    The nerve growth factor (NGF) is a neurotrophin regulating the survival and function peripheral sensory and sympathetic neurons and of forebrain cholinergic neurons. Both peripheral neuropathies and brain cholinergic dysfunctions could benefit from NGFbased therapies, but the clinical use of NGF has been so far hampered by the development of important side effects, like hyperalgesia and autonomic dysfunctions. Acupuncture is a therapeutic technique and is a part of traditional Chinese medicine. Western descriptions of the clinical efficacy of acupuncture on pain, inflammation, motor dysfunction, mood disorders, and seizures are based on the stimulation of several classes of sensory afferent fibers and the consequent activation of physiological processes similar to those resulting from physical exercise or deep massage. Recently, it has been shown that peripheral sensory stimulation by electroacupuncture (EA) could improve brain NGF availability and utilization, at the same time counteracting the major sideeffects induced by NGF administration. This review focuses on the emerging links between EA and NGF with special emphasis on the work carried out in the last decade in our laboratory, investigating the role of NGF as a mediator of EA effects in the central nervous system and as a modulator of sensory and autonomic activity.

  17. Herpesvirus-mediated systemic delivery of nerve growth factor.

    PubMed

    Wolfe, D; Goins, W F; Kaplan, T J; Capuano, S V; Fradette, J; Murphey-Corb, M; Robbins, P D; Cohen, J B; Glorioso, J C

    2001-01-01

    Sustained systemic dissemination of therapeutic proteins from peripheral sites is an attractive prospect for gene therapy applications. Replication-defective genomic herpes simplex virus type 1 (HSV-1) vectors were evaluated for their ability to express nerve growth factor (NGF) as a model gene product both locally and systemically. Intra-articular inoculation of NGF expression vectors in rabbits resulted in significant increases in joint lavage and blood plasma NGF that persisted for 1 year. A rhesus macaque injected intra-articularly displayed a comparable increase in plasma NGF for at least 6 months, at which time the serum NGF levels of this animal were sufficient to cause differentiation of PC12 cells in culture, but not to increase footpad epidermis innervation. Long-term reporter transgene expression was observed primarily in ligaments, a finding confirmed by direct inoculation of patellar ligament. Patellar ligament inoculation with a NGF vector resulted in elevated levels of circulating NGF similar to those observed following intra-articular vector delivery. These results represent the first demonstration of sustained systemic release of a transgene product using HSV vectors, raising the prospect of new applications for HSV-1 vectors in the treatment of systemic disease.

  18. Choline Acetyltransferase Activity in Striatum of Neonatal Rats Increased by Nerve Growth Factor

    NASA Astrophysics Data System (ADS)

    Mobley, William C.; Rutkowski, J. Lynn; Tennekoon, Gihan I.; Buchanan, Karen; Johnston, Michael V.

    1985-07-01

    Some neurodegenerative disorders may be caused by abnormal synthesis or utilization of trophic molecules required to support neuronal survival. A test of this hypothesis requires that trophic agents specific for the affected neurons be identified. Cholinergic neurons in the corpus striatum of neonatal rats were found to respond to intracerebroventricular administration of nerve growth factor with prominent, dose-dependent, selective increases in choline acetyltransferase activity. Cholinergic neurons in the basal forebrain also respond to nerve growth factor in this way. These actions of nerve growth factor may indicate its involvement in the normal function of forebrain cholinergic neurons as well as in neurodegenerative disorders involving such cells.

  19. A minority of carcinoma cells producing acidic fibroblast growth factor induces a community effect for tumor progression.

    PubMed Central

    Jouanneau, J; Moens, G; Bourgeois, Y; Poupon, M F; Thiery, J P

    1994-01-01

    It is generally accepted that primary tumors become heterogeneous as a consequence of tumor-cell genetic instability. Clonal dominance has been shown to occur in some experimental models allowing a subpopulation of cells to overgrow the primary heterogeneous tumor and to metastasize. Alternatively, interactions among coexisting tumor subpopulations may contribute to the emergence of a malignant invasive primary solid tumor. We asked the question whether emergence of carcinoma cells producing a growth/dissociating factor within a tumor cell population may be a determinant for tumor progression and for clonal dominance. To mimic such a situation, we have investigated the impact of tumor subpopulation heterogeneity in an in vivo model in which mixtures of carcinoma cells that differ in their ability to produce acidic fibroblast growth factor are injected into nude mice. Our data indicate that a growth-factor-producing cell subpopulation can confer increased tumorigenicity to an entire cell population and subsequently elicit a shorter delay for appearance of metastasis. A community effect via cell interactions may account for a heterogeneous tumor cell population rather than clonal dominance during progression of certain tumor types. Images Fig. 3 PMID:7506417

  20. Retrograde axonal transport of /sup 125/I-nerve growth factor in rat ileal mesenteric nerves. Effect of streptozocin diabetes

    SciTech Connect

    Schmidt, R.E.; Plurad, S.B.; Saffitz, J.E.; Grabau, G.G.; Yip, H.K.

    1985-12-01

    The retrograde axonal transport of intravenously (i.v.) administered /sup 125/I-nerve growth factor (/sup 125/I-NGF) was examined in mesenteric nerves innervating the small bowel of rats with streptozocin (STZ) diabetes using methods described in detail in the companion article. The accumulation of /sup 125/I-NGF distal to a ligature on the ileal mesenteric nerves of diabetic animals was 30-40% less than in control animals. The inhibition of accumulation of /sup 125/I-NGF in diabetic animals was greater at a ligature tied 2 h after i.v. administration than at a ligature tied after 14 h, which suggests that the diabetic animals may have a lag in initiation of NGF transport in the terminal axon or retardation of transport at some site along the axon. The /sup 125/I-NGF transport defect was observed as early as 3 days after the induction of diabetes, a time before the development of structural axonal lesions, and did not worsen at later times when dystrophic axonopathy is present. Both the ileal mesenteric nerves, which eventually develop dystrophic axonopathy in experimental diabetes, and the jejunal mesenteric nerves, which never develop comparable structural alterations, showed similar /sup 125/I-NGF transport deficits, suggesting that the existence of the transport abnormality does not predict the eventual development of dystrophic axonal lesions. Autoradiographic localization of /sup 125/I-NGF in the ileal mesenteric nerves of animals that had been diabetic for 11-13 mo demonstrated decreased amounts of /sup 125/I-NGF in transit in unligated paravascular nerve fascicles. There was, however, no evidence for focal retardation of transported /sup 125/I-NGF at the sites of dystrophic axonal lesions.

  1. Mechanisms for growth factor-induced pituitary tumor transforming gene-1 expression in pituitary folliculostellate TtT/GF cells.

    PubMed

    Vlotides, G; Cruz-Soto, M; Rubinek, T; Eigler, T; Auernhammer, C J; Melmed, S

    2006-12-01

    PTTG1, a securin protein, also behaves as a transforming gene and is overexpressed in pituitary tumors. Because pituitary folliculostellate (FS) cells regulate pituitary tumor growth factors by paracrine mechanisms, epidermal growth factor (EGF) receptor (EGFR)-mediated PTTG1 expression and cell proliferation was tested in pituitary FS TtT/GF cells. EGFR ligands caused up to 3-fold induction of Pttg1 mRNA expression, enhanced proliferating cell nuclear antigen, and increased entry of G0/1-arrested cells into S-phase. PTTG binding factor mRNA expression was not altered. EGF-induced Pttg1 expression and cell proliferation was abolished by preincubation of TtT/GF cells with EGFR inhibitors AG1478 and gefitinib. Phosphatidylinositol 3 kinase, protein kinase C, and MAPK, but not c-Jun N-terminal kinase and Janus activating kinase signaling regulated EGF-induced Pttg1, as well as proliferating cell nuclear antigen mRNA expression and entry into S-phase. EGF-induced EGFR and ERK1/2 phosphorylation was followed by rapid MAPK kinase/ERK kinase-dependent activation of Elk-1 and c-Fos. EGF-induced Pttg1 expression peaked at the S-G2 transition and declined thereafter. Pttg1 cell cycle dependency was confirmed by suppression of EGF-induced Pttg1 mRNA by blockade of cells in early S-phase. The results show that PTTG1 and its binding protein PTTG binding factor are expressed in pituitary FS TtT/GF cells. EGFR ligands induce PTTG1 and regulate S-phase, mediated by phosphatidylinositol 3 kinase, protein kinase C, and MAPK pathways. PTTG1 is therefore a target for EGFR-mediated paracrine regulation of pituitary cell growth.

  2. Microscopic analysis of the cellular events during scatter factor/hepatocyte growth factor-induced epithelial tubulogenesis.

    PubMed

    Williams, M J; Clark, P

    2003-11-01

    Scatter factor/hepatocyte growth factor (SF/HGF), a large multifunctional polypeptide growth and motility factor, is known to play important roles during embryonic development, adult tissue growth and repair. In an established three-dimensional type I collagen model, SF/HGF induces Madin-Darby canine kidney (MDCK) epithelial cysts to form long, branching tubules (tubulogenesis). In addition, the composition of the surrounding extracellular matrix (ECM) has been shown to modulate SF/HGF-induced morphogenesis, where tubulogenesis was completely abrogated in Matrigel basement membrane. Many cellular events that occur during SF/HGF-mediated remodelling, and its modulation by the ECM, remain unclear. We have investigated these mechanisms through microscopic examination of the time-course of SF/HGF-induced responses in MDCK cysts cultured in type I collagen or Matrigel. We found that early responses to SF/HGF were matrix-independent. Changes included increased paracellular spacing between normally closely apposed lateral membranes, and the formation of filopodial processes, indicating a partial motile response. Cell-cell contact was maintained, with the persistence of cell junctions. Therefore, while one or a number of ECM components are preventing SF/HGF-primed cells from undergoing an invasive and/or migratory programme, non-permissive matrices are not preventing SF/HGF signalling to the cell. Later matrix-dependent responses, which occurred in type I collagen but not Matrigel, included the formation of basal protrusions that comprise two or more neighbouring cells, which extend to form nascent tubules. Modified polarity of cells comprising the basal protrusions was evident, with a marker for the apical membrane being found in the same region as adherens junctions and desmosomes, typically localized at lateral membranes. We propose a model for SF/HGF-induced tubulogenesis in which tubules form from basal protrusions of adjacent cells. This mechanism of in vitro tubule

  3. The Glucuronyltransferase GlcAT-P Is Required for Stretch Growth of Peripheral Nerves in Drosophila

    PubMed Central

    Pandey, Rahul; Blanco, Jorge; Udolph, Gerald

    2011-01-01

    During development, the growth of the animal body is accompanied by a concomitant elongation of the peripheral nerves, which requires the elongation of integrated nerve fibers and the axons projecting therein. Although this process is of fundamental importance to almost all organisms of the animal kingdom, very little is known about the mechanisms regulating this process. Here, we describe the identification and characterization of novel mutant alleles of GlcAT-P, the Drosophila ortholog of the mammalian glucuronyltransferase b3gat1. GlcAT-P mutants reveal shorter larval peripheral nerves and an elongated ventral nerve cord (VNC). We show that GlcAT-P is expressed in a subset of neurons in the central brain hemispheres, in some motoneurons of the ventral nerve cord as well as in central and peripheral nerve glia. We demonstrate that in GlcAT-P mutants the VNC is under tension of shorter peripheral nerves suggesting that the VNC elongates as a consequence of tension imparted by retarded peripheral nerve growth during larval development. We also provide evidence that for growth of peripheral nerve fibers GlcAT-P is critically required in hemocytes; however, glial cells are also important in this process. The glial specific repo gene acts as a modifier of GlcAT-P and loss or reduction of repo function in a GlcAT-P mutant background enhances VNC elongation. We propose a model in which hemocytes are required for aspects of glial cell biology which in turn affects the elongation of peripheral nerves during larval development. Our data also identifies GlcAT-P as a first candidate gene involved in growth of integrated peripheral nerves and therefore establishes Drosophila as an amenable in-vivo model system to study this process at the cellular and molecular level in more detail. PMID:22132223

  4. Influence of congenital facial nerve palsy on craniofacial growth in craniofacial microsomia.

    PubMed

    Choi, Jaehoon; Park, Sang Woo; Kwon, Geun-Yong; Kim, Sang-Hyun; Hur, Ji An; Baek, Seung-Hak; Kim, Jae Chan; Choi, Tae Hyun; Kim, Sukwha

    2014-11-01

    Facial muscles are of major importance in human craniofacial growth and development. The purpose of our study was to investigate whether congenital facial nerve palsy influences craniofacial growth in craniofacial microsomia. Fifty-one patients with unilateral craniofacial microsomia and no history of craniofacial skeletal surgery whose radiographs were taken after craniofacial growth was complete were included in this study. These patients were divided into groups in which the facial nerve was involved or uninvolved. The authors evaluated a total of seven measurement items to analyze the midface and mandibular asymmetry. Twenty patients had facial nerve involvement, and 31 had no involvement. None of the measurement items revealed any significant differences between the facial nerve-involved group and the uninvolved group within the same modified Pruzansky grade. There was no correlation between the type of facial nerve involvement and the measurement items. In relationships among the measurement items within each group, maxillary asymmetry was indirectly correlated with mandibular asymmetry or midline deviation through the occlusal plane angle in the uninvolved groups. However, in the facial nerve-involved group, the relationships disappeared. When the correlations in the facial nerve-involved group were compared with those of the uninvolved group, the relationships in the uninvolved group appeared more significant than in the facial nerve-involved group. The loss of relationships between the upper and lower jaw in the facial nerve-involved group might have been caused by subtle changes, which occur in midfacial bones and in the mandible due to facial nerve palsy. The main limitation of our study is that aside from facial nerve palsy, craniofacial microsomia has many factors that can influence craniofacial growth, such as hypoplasia of the mandibular condyle and soft tissue deficiencies. Copyright © 2014 British Association of Plastic, Reconstructive and

  5. Nerve growth factor and associated nerve sprouting contribute to local mechanical hyperalgesia in a rat model of bone injury.

    PubMed

    Yasui, M; Shiraishi, Y; Ozaki, N; Hayashi, K; Hori, K; Ichiyanagi, M; Sugiura, Y

    2012-08-01

    To clarify the mechanism of tenderness after bone injury, we investigated changes in the withdrawal threshold to mechanical stimuli, nerve distribution and nerve growth factor (NGF)-expression in a rat model of bone injury without immobilization for bone injury healing. Rats were divided into three groups as follows: (1) rats incised in the skin and periosteum, followed by drilling a hole in the tibia [bone lesion group (BLG)]; (2) those incised in the skin and periosteum without bone drilling [periosteum lesion group (PLG)]; and (3) those incised in the skin [skin lesion group (SLG)]. Mechanical hyperalgesia continued for 28 days at a lesion in the BLG, 21 days in PLG and 5 days in SLG after treatments, respectively. Endochondral ossification was observed on days 5-28 in BLG and on days 5-21 in PLG. Nerve growth appeared in deep connective tissue (DCT) at day 28 in BLG. Nerve fibres increased in both cutaneous tissue and DCT at day 7 in PLG, but they were not found at day 28. Mechanical hyperalgesia accompanied with endochondral ossification and nerve fibres increasing at the lesion in both BLG and PLG. NGF was expressed in bone-regenerating cells during the bone injury healing. Anti-NGF and trk inhibitor K252a inhibited hyperalgesia in the different time course. This study shows that localized tenderness coincides with the bone healing and involves NGF expression and nerve sprouting after bone injury. The findings present underlying mechanisms and provide pathophysiological relevance of local tenderness to determination of bone fracture and its healing.

  6. A quantitative proteomic analysis of growth factor-induced compositional changes in lipid rafts of human smooth muscle cells.

    PubMed

    MacLellan, Dawn L; Steen, Hanno; Adam, Rosalyn M; Garlick, Monica; Zurakowski, David; Gygi, Steven P; Freeman, Michael R; Solomon, Keith R

    2005-12-01

    Signals that promote proliferation and migration of smooth muscle cells (SMC) have been implicated in pathologic growth of hollow organs. Members of the platelet-derived growth factor (PDGF) family, potent mitogens and motility factors for SMC, have been shown to signal through cholesterol-enriched lipid rafts. We recently demonstrated that PDGF-stimulated DNA synthesis in urinary tract SMC was dependent on the integrity of lipid rafts. Despite its known ability to rapidly alter discrete proteins within rafts, the effect of PDGF on overall raft protein composition is unknown. In this study, we employed isotope coded affinity tag (ICAT) analysis to evaluate PDGF-induced protein changes in lipid rafts of primary culture human SMC. Following acute (i.e., 15 min) exposure of SMC to PDGF, 23 proteins increased in rafts >20%. In contrast, raft localization of only three proteins increased after 12 h of PDGF treatment. Among the proteins that increased at 15 min were the glycophosphatidylinositol-anchored proteins Thy-1, 5'-nucleotidase, and CD55, the cytoskeletal proteins actin, actinin, tropomyosin-3 and -4, and the endocytosis-related proteins clathrin and beta-adaptin. In addition, eight Rho family members were localized to rafts by ICAT analysis. Collectively, these observations suggest a role for lipid rafts in regulation of PDGF-stimulated changes in the cytoskeleton.

  7. Nerve growth factor in human semen: Effect of nerve growth factor on the normozoospermic men during cryopreservation process

    PubMed Central

    Saeednia, Sara; Bahadoran, Hosein; Amidi, Fardin; Asadi, Mohammad Hosein; Naji, Mohammad; Fallahi, Parvin; Nejad, Nahid Ataie

    2015-01-01

    Objective(s): Although routinely applied in assisted reproductive technology, human sperm cryopreservation is not a completely successful procedure. Adverse effects of cryopreservation on the fertilization capacity, motility, morphology, and viability of spermatozoa have been proven; cryopreservation has also shown a role in sperm DNA fragmentation and infertility. The post-thaw survival of spermatozoa improved after addition of supplementation of antioxidant molecules to freezing media. Nerve growth factor (NGF) as one of the prosurvival substances has gained great attention in recent years. The aim of this study was the usage of NGF as prosurvival factor after cryopreservation process of human semen samples to assess the motility and viability of sperm, nitric oxide (NO) concentration, and DNA fragmentation in normozoospermic men. Materials and Methods: Semen samples were collected from 25 normozoospermic men and were divided into fresh semen samples as control group, frozen–thawed semen samples without addition of exogenous NGF, and three groups of semen samples cryopreserved with addition of exogenous NGF (0.5, 1, and 5 ng/ml) in freezing medium. Viability was assessed by eosin-negrosin staining technique. Motility was evaluated with inverted microscope. NO concentration and apoptosis content were measured with flow cytometry. Results: Results showed that exogenous NGF at 0.5 ng/ml could significantly (P-value <0.05) influence viability, motility, nitric oxide, and DNA fragmentation content. Conclusion: Exogenous NGF as cryoprotectant improved sperm viability and motility, increased intracellular NO concentration, and decreased apoptosis content in normal human spermatozoa. PMID:25945243

  8. ErbB4 isoforms selectively regulate growth factor induced Madin-Darby canine kidney cell tubulogenesis.

    PubMed

    Zeng, Fenghua; Zhang, Ming-Zhi; Singh, Amar B; Zent, Roy; Harris, Raymond C

    2007-11-01

    ErbB4, a member of the epidermal growth factor (EGF) receptor family that can be activated by heregulin beta1 and heparin binding (HB)-EGF, is expressed as alternatively spliced isoforms characterized by variant extracellular juxtamembrane (JM) and intracellular cytoplasmic (CYT) domains. ErbB4 plays a critical role in cardiac and neural development. We demonstrated that ErbB4 is expressed in the ureteric buds and developing tubules of embryonic rat kidney and in collecting ducts in adult. The predominant isoforms expressed in kidney are JM-a and CYT-2. In ErbB4-transfected MDCK II cells, basal cell proliferation and hepatocyte growth factor (HGF)-induced tubule formation were decreased by all four isoforms. Only JM-a/CYT-2 cells formed tubules upon HB-EGF stimulation. ErbB4 was activated by both HRG-beta1 and HB-EGF stimulation; however, compared with HRG-beta1, HB-EGF induced phosphorylation of the 80-kDa cytoplasmic cleavage fragment of the JM-a/CYT-2 isoform. HB-EGF also induced early activation of ERK1/2 in JM-a/CYT-2 cells and promoted nuclear translocation of the JM-a/CYT-2 cytoplasmic tail. In summary, our data indicate that JM-a/CYT-2, the ErbB4 isoform that is proteinase cleavable but does not contain a PI3K-binding domain in its cytoplasmic tail, mediates important functions in renal epithelial cells in response to HB-EGF.

  9. Mechanisms in decorin regulation of vascular endothelial growth factor-induced human trophoblast migration and acquisition of endothelial phenotype.

    PubMed

    Lala, Neena; Girish, Gannareddy V; Cloutier-Bosworth, Alia; Lala, Peeyush K

    2012-09-01

    Extravillous trophoblast (EVT) cells of the human placenta invade the uterine decidua and utero-placental arteries to establish an efficient exchange of key molecules between maternal and fetal blood. Trophoblast invasion is stringently regulated in situ both positively and negatively by a variety of factors at the fetal-maternal interface to maintain a healthy utero-placental homeostasis. One such factor, decorin, a transforming growth factor (TGF)-beta binding, leucine-rich proteoglycan produced by the decidua, negatively regulates EVT proliferation, migration, and invasiveness independent of TGF-beta. We reported that these decorin actions were mediated by its binding to multiple tyrosine kinase receptors, including vascular endothelial growth factor receptor (VEGFR)-2. The present study explores the mechanisms underlying decorin antagonism of VEGF (VEGF-A) stimulation of endovascular differentiation of EVT using our EVT cell line, HTR-8/SVneo. We observe that decorin inhibits VEGF-induced EVT cell migration and endothelial-like tube formation on matrigel. VEGF activates MAPKs (p38 MAPK, MEK3/6, and ERK1/2) in EVT cells, and the activation is blocked in both cases by decorin. Employing selective MAPK inhibitors, we show that both p38 and ERK pathways contribute independently to VEGF-induced EVT migration and capillary-like tube formation. VEGF upregulates the vascular endothelial (VE) markers VE-cadherin and beta-catenin in EVT and endothelial cells, and this upregulation is blocked by decorin and MAPK inhibitors. These results suggest that decorin inhibits VEGF-A stimulation of trophoblast migration and endovascular differentiation by interfering with p38 MAPK and ERK1/2 activation. Thus decorin-mediated dual impediment of endovascular differentiation of the EVT and angiogenesis may have implications for pathogenesis of preeclampsia, a hypoinvasive trophoblast disorder in pregnancy.

  10. Sphingosine Kinase Mediates Vascular Endothelial Growth Factor-Induced Activation of Ras and Mitogen-Activated Protein Kinases

    PubMed Central

    Shu, Xiaodong; Wu, Weicheng; Mosteller, Raymond D.; Broek, Daniel

    2002-01-01

    Vascular endothelial growth factor (VEGF) signaling is critical to the processes of angiogenesis and tumor growth. Here, evidence is presented for VEGF stimulation of sphingosine kinase (SPK) that affects not only endothelial cell signaling but also tumor cells expressing VEGF receptors. VEGF or phorbol 12-myristate 13-acetate treatment of the T24 bladder tumor cell line resulted in a time- and dose-dependent stimulation of SPK activity. In T24 cells, VEGF treatment reduced cellular sphingosine levels while raising that of sphingosine-1-phosphate. VEGF stimulation of T24 cells caused a slow and sustained accumulation of Ras-GTP and phosphorylated extracellular signal-regulated kinase (phospho-ERK) compared with that after EGF treatment. Small interfering RNA (siRNA) that targets SPK1, but not SPK2, blocks VEGF-induced accumulation of Ras-GTP and phospho-ERK in T24 cells. In contrast to EGF stimulation, VEGF stimulation of ERK1/2 phosphorylation was unaffected by dominant-negative Ras-N17. Raf kinase inhibition blocked both VEGF- and EGF-stimulated accumulation of phospho-ERK1/2. Inhibition of SPK by pharmacological inhibitors, a dominant-negative SPK mutant, or siRNA that targets SPK blocked VEGF, but not EGF, induction of phospho-ERK1/2. We conclude that VEGF induces DNA synthesis in a pathway which sequentially involves protein kinase C (PKC), SPK, Ras, Raf, and ERK1/2. These data highlight a novel mechanism by which SPK mediates signaling from PKC to Ras in a manner independent of Ras-guanine nucleotide exchange factor. PMID:12391145

  11. Liver growth factor induces testicular regeneration in EDS-treated rats and increases protein levels of class B scavenger receptors.

    PubMed

    Lobo, M V T; Arenas, M I; Huerta, L; Sacristán, S; Pérez-Crespo, M; Gutiérrez-Adán, A; Díaz-Gil, J J; Lasunción, M A; Martín-Hidalgo, A

    2015-01-15

    The aim of the present work was to determine the effects of liver growth factor (LGF) on the regeneration process of rat testes after chemical castration induced by ethane dimethanesulfonate (EDS) by analyzing some of the most relevant proteins involved in cholesterol metabolism, such as hormone sensitive lipase (HSL), 3β-hydroxysteroid dehydrogenase (3β-HSD), scavenger receptor SR-BI, and other components of the SR family that could contribute to the recovery of steroidogenesis and spermatogenesis in the testis. Sixty male rats were randomized to nontreated (controls) and LGF-treated, EDS-treated, and EDS + LGF-treated groups. Testes were obtained on days 10 (T1), 21 (T2), and 35 (T3) after EDS treatment, embedded in paraffin, and analyzed by immunohistochemistry and Western blot. LGF improved the recovery of the seminiferous epithelia, the appearance of the mature pattern of Leydig cell interstitial distribution, and the expression of mature SR-BI. Moreover, LGF treatment resulted in partial recovery of HSL expression in Leydig cells and spermatogonia. No changes in serum testosterone were observed in control or LGF-treated rats, but in EDS-castrated animals LGF treatment induced a progressive increase in serum testosterone levels and 3β-HSD expression. Based on the pivotal role of SR-BI in the uptake of cholesteryl esters from HDL, it is suggested that the observed effects of LGF would facilitate the provision of cholesterol for sperm cell growth and Leydig cell recovery. Copyright © 2015 the American Physiological Society.

  12. Coordinated Movement of Vesicles and Actin Bundles during Nerve Growth Revealed by Superresolution Microscopy.

    PubMed

    Nozumi, Motohiro; Nakatsu, Fubito; Katoh, Kaoru; Igarashi, Michihiro

    2017-02-28

    The growth cone is an essential structure for nerve growth. Although its membrane and cytoskeleton are likely to interact coordinately during nerve growth, the mechanisms are unknown due to their close proximity. Here, we used superresolution microscopy to simultaneously observe vesicles and F-actin in growth cones. We identified a novel vesicular generation mechanism that is independent of clathrin and dependent on endophilin-3- and dynamin-1 and that occurs proximal to the leading edge simultaneously with fascin-1-dependent F-actin bundling. In contrast to conventional clathrin-dependent endocytosis, which occurs distal from the leading edge at the basal surfaces of growth cones, this mechanism was distinctly observed at the apical surface using 3D imaging and was involved in mediating axon growth. Reduced endophilin or fascin inhibited this endocytic mechanism. These results suggest that, at the leading edge, vesicles are coordinately generated and transported with actin bundling during nerve growth.

  13. Can amino-functionalized carbon nanotubes carry functional nerve growth factor?

    PubMed Central

    Chen, Wen; Xiong, Qing; Ren, Quanxia; Guo, Yake; Li, Gao

    2014-01-01

    Carbon nanotubes can carry protein into cells to induce biological effects. Amino-functionalized carbon nanotubes are soluble and biocompatible, have high reactivity and low toxicity, and can help promote nerve cell growth. In this study, amino-functionalized ethylenediamine-treated multi-walled carbon nanotubes were used to prepare carbon nanotubes-nerve growth factor complexes by non-covalent grafting. The physicochemical properties, cytotoxicity to PC12 and chick embryo dorsal root ganglion, and biological activity of the carbon nanotubes-nerve growth factor complexes were investigated. The results showed that amino functionalization improved carbon nanotubes-nerve growth factor complex dispersibility, reduced their toxicity to PC12 cells, and promoted PC12 cell differentiation and chick embryo dorsal root ganglion. PMID:25206814

  14. Adipose-derived stem cells stimulate regeneration of peripheral nerves: BDNF secreted by these cells promotes nerve healing and axon growth de novo.

    PubMed

    Lopatina, Tatiana; Kalinina, Natalia; Karagyaur, Maxim; Stambolsky, Dmitry; Rubina, Kseniya; Revischin, Alexander; Pavlova, Galina; Parfyonova, Yelena; Tkachuk, Vsevolod

    2011-03-14

    Transplantation of adipose-derived mesenchymal stem cells (ASCs) induces tissue regeneration by accelerating the growth of blood vessels and nerve. However, mechanisms by which they accelerate the growth of nerve fibers are only partially understood. We used transplantation of ASCs with subcutaneous matrigel implants (well-known in vivo model of angiogenesis) and model of mice limb reinnervation to check the influence of ASC on nerve growth. Here we show that ASCs stimulate the regeneration of nerves in innervated mice's limbs and induce axon growth in subcutaneous matrigel implants. To investigate the mechanism of this action we analyzed different properties of these cells and showed that they express numerous genes of neurotrophins and extracellular matrix proteins required for the nerve growth and myelination. Induction of neural differentiation of ASCs enhances production of brain-derived neurotrophic factor (BDNF) as well as ability of these cells to induce nerve fiber growth. BDNF neutralizing antibodies abrogated the stimulatory effects of ASCs on the growth of nerve sprouts. These data suggest that ASCs induce nerve repair and growth via BDNF production. This stimulatory effect can be further enhanced by culturing the cells in neural differentiation medium prior to transplantation.

  15. Distinct growth factor-induced dynamic mass redistribution (DMR) profiles for monitoring oncogenic signaling pathways in various cancer cells.

    PubMed

    Du, Yuhong; Li, Zijian; Li, Lian; Chen, Zhuo Georgia; Sun, Shi-Yong; Chen, Peifang; Shin, Dong M; Khuri, Fadlo R; Fu, Haian

    2009-01-01

    Targeting dysregulated signaling pathways in tumors has led to the development of a novel class of signal transduction inhibitors, including inhibitors of the epidermal growth factor (EGF) receptor (EGFR). To dissect oncogenic pathways, identify key pathway determinants, and evaluate the efficacy of targeted agents, it is vital to develop technologies that allow the detection of temporal signaling events under physiological conditions. Here we report the application of a label-free optical biosensor to reveal the rapid response of cancer cells to EGF, expressed as a dynamic mass redistribution (DMR) signal. In response to EGF, squamous cell carcinoma of the head and neck cells exhibited a rapid rise in DMR signal, whereas lung adenocarcinoma cells showed a biphasic DMR profile, suggesting a cell type-dependent DMR response. Pharmacological studies suggested the importance of EGFR and the phosphatidylinositol-3 kinase pathway in mediating the EGF-induced DMR response. The defined DMR signatures offer a simple yet sensitive tool for evaluating EGFR-targeted agents, as shown with gefitinib and erlotinib. The assay can also be used for cell-based high-throughput screening of EGF pathway inhibitors, as demonstrated by its robust performance in a 384-well plate format (Z' > 0.5). This technology is applicable to other oncogenic pathways for the discovery of novel therapeutic agents for the treatment of various cancers.

  16. Demonstration of inhibitory effect of oral shark cartilage on basic fibroblast growth factor-induced angiogenesis in the rabbit cornea.

    PubMed

    González, R P; Soares, F S; Farias, R F; Pessoa, C; Leyva, A; de Barros Viana, G S; Moraes, M O

    2001-02-01

    Several angiogenic inhibitors have been obtained from shark cartilage, some of these are currently in clinical trials for assessment of safety and therapeutic efficacy in humans. Still, shark cartilage taken orally is commonly used in alternative and complimentary medicine for various ailments including serious diseases such as cancer. However, only few studies of oral shark cartilage have demonstrated pharmacological effects in experimental animals or patients, to indicate safe doses with sufficient bioavailability. In the present study we demonstrated the antiangiogenic properties of oral shark cartilage in the rabbit cornea model. Slow-release, polymethylmetacrylate pellets containing basic fibroblast growth factor (bFGF) were surgically implanted in the rabbit cornea to stimulate neovascularization scored by stereo microscopy. Powdered shark cartilage (PSC; commercial product) was tested orally along with a water-soluble fraction (WSF) of this cartilage product which was tested by local application. Animals were treated with oral dosages of 100 mg/kg PSC or 200 mg/kg thalidomide as positive control. Pellets containing WSF (50, 100 or 200 microg/pellet) or bFGF-inhibitor pentosan polysulfate were implanted adjacent to the bFGF pellet. Oral shark cartilage inhibited bFGF-induced angiogenesis, as did oral thalidomide, in this in vivo model. WSF and pentosan polysulfate was shown to block neovascularization in the cornea when applied locally. This study demonstrates that in the rabbit, oral shark cartilage appears to produce systemic levels of angiogenesis inhibitors that can exert their effect at the cornea.

  17. Critical role of aquaporin-3 in epidermal growth factor-induced migration of colorectal carcinoma cells and its clinical significance.

    PubMed

    Li, Ang; Lu, Dehong; Zhang, Yupeng; Li, Jia; Fang, Yu; Li, Fei; Sun, Jiabang

    2013-02-01

    Aquaporins (AQPs) are a family of small, integral membrane proteins that have been shown to play an important role in tumor development and metastasis. Several studies have demonstrated that expression of AQP3 contributes to the enhanced migration of epithelial cells and is related to differentiation, metastasis and vascular invasion in lung and gastric cancer. Therefore, we investigated whether AQP3 could enhance human colorectal carcinoma cell migration and we examined the role of AQP3 in the prognosis of colorectal carcinoma. Our results showed that human epidermal growth factor (hEGF) increased the expression of AQP3 and, subsequently, the migration ability of human colorectal carcinoma cells HCT116 in a dose- and time-dependent manner. The enhanced migration ability of HCT116 cells was blocked by the AQP3 inhibitor, CuSO(4). Overexpression of AQP3 induced by hEGF was inhibited by a PI3K/AKT inhibitor, LY294002, but the ERK inhibitor U0126 had a minor effect on the hEGF-induced AQP3 upregulation. Immunohistochemical staining of the cancer tissues and corresponding normal tissues showed that AQP3 expression in cancer tissue was higher compared to that in normal tissue. The expression intensity of AQP3 was associated with the differentiation, lymph node and distant metastasis of colorectal carcinoma patients. Our results suggest that AQP3 overexpression could facilitate colorectal carcinoma cell migration and AQP3 may be considered a potential indicator and therapeutic target for colon tumor metastasis and prognosis.

  18. Epidermal growth factor-induced mobilization of a ganglioside-specific sialidase (NEU3) to membrane ruffles

    SciTech Connect

    Yamaguchi, Kazunori; Hata, Keiko; Wada, Tadashi; Moriya, Setsuko; Miyagi, Taeko . E-mail: miyagi-ta173@pref.miyagi.jp

    2006-07-28

    Human ganglioside-specific sialidase, NEU3, localized at cell membranes is thought to regulate various biological processes at cell surfaces. We here explored functional subcellular localization of the sialidase by immunofluorescence and found accumulation at leading edges of cell membranes in the presence of serum in culture. In response to EGF, the sialidase redistributed rapidly to ruffling cell membranes of squamous carcinoma A431 cells and co-localized with Rac-1. NEU3 overexpression enhanced Rac-1 activation and cell migration as compared with controls in HeLa cells as well as in A431 cells. Consistent with co-localization with Rac-1 by immunofluorescence, NEU3 was found to co-precipitate with activated Rac bound to GST-PAK-1 fusion protein. NEU3 silencing by siRNA, in contrast, resulted in inhibition of Rac-1 activation. These results indicate that NEU3 is able to mobilize to membrane ruffles in response to growth stimuli and activate the Rac-1 signaling by co-localization with Rac-1, leading to increased cell motility.

  19. MET inhibitor PHA-665752 suppresses the hepatocyte growth factor-induced cell proliferation and radioresistance in nasopharyngeal carcinoma cells

    SciTech Connect

    Liu, Tongxin; Li, Qi; Sun, Quanquan; Zhang, Yuqin; Yang, Hua; Wang, Rong; Chen, Longhua; Wang, Wei

    2014-06-20

    Highlights: • We demonstrated that irradiation induced MET overexpression and activation. • The aberrant MET signal mediated by HGF induced proliferation and radioresistance of NPC cells. • MET inhibitor PHA-665752 effectively suppressed HGF induced cell proliferation and radioresistance in NPC cells. • PHA-665752 suppressed the three downstream pathway of HGF/MET signal in a dose-dependent manner. - Abstract: Although ionizing radiation (IR) has provided considerable improvements in nasopharyngeal carcinoma (NPC), in subsets of patients, radioresistance is still a major problem in the treatment. In this study, we demonstrated that irradiation induced MET overexpression and activation, and the aberrant MET signal mediated by hepatocyte growth factor (HGF) induced radioresistance. We also found that MET inhibitor PHA-665752 effectively suppressed HGF induced cell proliferation and radioresistance in NPC cells. Further investigation indicated that PHA-665752 suppressed the phosphorylation of the Akt, ERK1/2, and STAT3 proteins in a dose-dependent manner. Our data indicated that the combination of IR with a MET inhibitor, such as PHA-665752, might be a promising therapeutic strategy for NPC.

  20. Temporal mismatch between pain behaviour, skin Nerve Growth Factor and intra-epidermal nerve fibre density in trigeminal neuropathic pain

    PubMed Central

    2014-01-01

    Background The neurotrophin Nerve Growth factor (NGF) is known to influence the phenotype of mature nociceptors, for example by altering synthesis of neuropeptides, and changes in NGF levels have been implicated in the pathophysiology of chronic pain conditions such as neuropathic pain. We have tested the hypothesis that after partial nerve injury, NGF accumulates within the skin and causes ‘pro-nociceptive’ phenotypic changes in the remaining population of sensory nerve fibres, which could underpin the development of neuropathic pain. Results Eleven days after chronic constriction injury of the rat mental nerve the intra-epidermal nerve fibre density of the chin skin from had reduced from 11.6 ± 4.9 fibres/mm to 1.0 ± 0.4 fibres/mm; this slowly recovered to 2.4 ± 2.0 fibres/mm on day 14 and 4.0 ± 0.8 fibres/mm on day 21. Cold hyperalgesia in the ipsilateral lower lip was detectable 11 days after chronic constriction injury, although at this time skin [NGF] did not differ between sides. At 14 days post-injury, there was a significantly greater [NGF] ipsilaterally compared to contralaterally (ipsilateral = 111 ± 23 pg/mg, contralateral = 69 ± 13 pg/mg), but there was no behavioural evidence of neuropathic pain at this time-point. By 21 days post-injury, skin [NGF] was elevated bilaterally and there was a significant increase in the proportion of TrkA-positive (the high-affinity NGF receptor) intra-epidermal nerve fibres that were immunolabelled for the neuropeptide Calcitonin Gene-related peptide. Conclusions The temporal mismatch in behaviour, skin [NGF] and phenotypic changes in sensory nerve fibres indicate that increased [NGF] does not cause hyperalgesia after partial mental nerve injury, although it may contribute to the altered neurochemistry of cutaneous nerve fibres. PMID:24380503

  1. Nerve growth factor protects against aluminum-mediated cell death.

    PubMed

    Ohyashiki, Takao; Satoh, Eiko; Okada, Morihiro; Takadera, Tsuneo; Sahara, Masako

    2002-07-15

    In the present study, we examined the effect of two salts of aluminum (Al), aluminum maltolate (Almal) and aluminum chloride (AlCl(3)), on the cell viability of PC12 cells in the absence and presence of nerve growth factor (NGF). A 72-h exposure of PC12 cells to Almal (300 microM) resulted in a marked increase of lactic dehydrogenase (LDH) release from the cells and a decrease of 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) activity. These results indicate that Almal induces a decrease in the cell viability. Under the same conditions, Almal also caused DNA ladder formation and chromatin condensation. In contrast, AlCl(3) did not showed an increased LDH release and a decreased MTT activity in the concentration range of the salt tested (0.1-1 mM). The extent of LDH release and MTT activity decrease induced by Almal treatment closely depended on the amount of Almal incorporated into the cells. An increase in the fluorescence intensity of 6-carboxy-2',7'-dichlorodihydrofluorescein diacetate, di(acetoxymethyl ester) (C-DCDHF-DA) which was loaded into the cell by Almal treatment and its prevention by pyrrolodine dithiocarbamate, a potent antioxidant, suggested that Almal-induced cell death partly proceeds via reactive oxygen species (ROS) production. NGF effectively inhibited the increase of LDH release and the decrease of MTT activity, as well as DNA fragmentation and chromatin condensation. However, NGF did not inhibit the increase of C-DCDHF-DA fluorescence in the cells induced by Almal treatment. From these results, it is suggested that ROS production associated with accumulation of Al is one possible important factor in the onset of Al neurotoxicity via apoptotic cell death and that NGF protects against cell degeneration associated with Al accumulation, but independently of ROS production.

  2. Role of Urothelial Nerve Growth Factor in Human Bladder Function

    PubMed Central

    Birder, Lori A.; Wolf-Johnston, Amanda; Griffiths, Derek; Resnick, Neil M.

    2011-01-01

    Aims To test whether nerve growth factor (NGF) concentration in human bladder urothelium/suburothelium is related to detrusor overactivity (DO), bladder sensation, detrusor contractility, or other aspects of lower urinary tract function. Materials and Methods Concentration of NGF was measured (using ELISA) in superficial bladder biopsies from 27 women (mean age 52 years, range 22–82) after comprehensive videourodynamics and bladder diary. Approximately half (12/27) showed clear DO and half did not. Results There was no evidence for increased NGF concentration in subjects with DO (association negative by Mann–Whitney test, P = 0.23). NGF was not significantly associated with two measures of detrusor contractility (Spearman’s r = −0.29, P = 0.17; r = −0.20, P = 0.33); nor with four measures inversely related to sensation: volume at strong desire to void and maximum capacity on cystometry (r = −0.13, P = 0.53; r = −0.23, P = 0.28), and maximum voided volume and mean daytime voided volume on bladder diary (r = −0.29, P = 0.16; r = −0.16, P = 0.44). It was significantly associated with 24-hr urine output on bladder diary (Spearman’s r = −0.55, P = 0.004). Conclusions Elevated NGF levels in human urothelium/suburothelium are not strongly associated with DO, detrusor contractility or increased bladder sensation. NGF levels are lower in subjects with higher 24-hr urine output. This observation is consistent with a role for NGF in an active process (trafficking) involved in bladder filling. PMID:17266135

  3. Beta-nerve growth factor levels in newborn cord sera.

    PubMed

    Haddad, J; Vilge, V; Juif, J G; Maitre, M; Donato, L; Messer, J; Mark, J

    1994-06-01

    This study was designed to examine beta-nerve growth factor (NGF) levels in human cord blood by a two-site enzyme immunoassay using MAb 27/21 to mouse NGF and to determine whether beta-NGF levels show developmental changes. Blood was collected at delivery from 61 newborns, 55 neonates appropriate for gestational age (46 term infants and 9 premature infants), 5 neonates small for gestational age, and 1 neonate with congenital hydrocephalus. In addition, samples were collected from 2 microcephalic children (microcephaly vera) aged 15 and 18 mo, 2 control children, and 4 healthy adults. Mean levels of NGF in preterm infants (n = 9; 13.7 +/- 8 pg/mL) were significantly lower than levels in term infants (n = 47; 21.2 +/- 8.8 pg/mL; p = 0.034 by Mann-Whitney U test). There was no correlation between birth weight, length, head circumference, and beta-NGF levels. In microcephalic children, NGF levels were low (8 pg/mL) compared with control infants' values (22 pg/mL). In adults, beta-NGF levels were higher and ranged between 238 and 292 pg/mL. Our study demonstrates that beta-NGF levels can be assessed in human newborn sera using a two-site enzyme immunoassay with MAb 27/21 to mouse beta-NGF, that beta-NGF levels are extremely low in newborns compared with adults, that beta-NGF levels seems to show developmental changes, and that beta-NGF levels may be used to assess NGF utilization under normal and pathologic conditions such as cerebral malformations.

  4. Mental disorders, functional impairment, and nerve growth factor

    PubMed Central

    Salles, Fanny Helena Martins; Soares, Pedro San Martin; Wiener, Carolina David; Mondin, Thaise Campos; da Silva, Paula Moraes; Jansen, Karen; de Mattos Souza, Luciano Dias; da Silva, Ricardo Azevedo; Oses, Jean Pierre

    2017-01-01

    Nerve growth factor (NGF) is an important member of the neurotrophin family and its alteration has been associated with psychiatric disorders. Functionality consists of the activities that an individual can perform, as well as their social participation, which is an important factor in analyzing the carrier living conditions of subjects with psychiatric suffering. Several studies have evaluated functionality in bipolar disorder; however, no studies have evaluated the functionality in other mental disorders. There are also few studies investigating the association between functionality and the biological bases of mental disorders. This study aimed to evaluate the serum NGF levels in psychiatric patients and to verify a possible association between the serum neurotrophic levels and functionality. This was a cross-sectional study with a convenient sample obtained from the Public Mental Health Service from the south of Brazil. The final sample was composed of 286 patients enrolled from July 2013 to October 2014. Data was collected using a sociodemographic questionnaire, and the diagnosis was confirmed using the Mini International Neuropsychiatric Interview (M.I.N.I) and a Functioning Assessment Short Test. The serum NGF levels were determined using the enzyme-linked immunosorbent assay method. Statistical analyses were performed using IBM SPSS Statistic 21.0 software. NGF serum levels were increased significantly in patients with obsessive–compulsive disorder compared with patients with no obsessive–compulsive disorder (P=0.015). An increase in serum NGF levels in generalized anxiety disorder patients was observed compared with patients with no generalized anxiety disorder (P=0.047). NGF was negatively associated with autonomy (P=0.024, r=−0.136), work (P=0.040, r=−0.124), and cognition (P=0.024, r=−0.137), thereby showing that changes in serum levels of NGF are associated with functionality in mental disorders. PMID:28053561

  5. Developmental regulation of human truncated nerve growth factor receptor

    SciTech Connect

    DiStefano, P.S.; Clagett-Dame, M.; Chelsea, D.M.; Loy, R. )

    1991-01-01

    Monoclonal antibodies (designated XIF1 and IIIG5) recognizing distinct epitopes of the human truncated nerve growth factor receptor (NGF-Rt) were used in a two-site radiometric immunosorbent assay to monitor levels of NGF-Rt in human urine as a function of age. Urine samples were collected from 70 neurologically normal subjects ranging in age from 1 month to 68 years. By using this sensitive two-site radiometric immunosorbent assay, NGF-Rt levels were found to be highest in urine from 1-month old subjects. By 2.5 months, NGF-Rt values were half of those seen at 1 month and decreased more gradually between 0.5 and 15 years. Between 15 and 68 years, urine NGF-Rt levels were relatively constant at 5% of 1-month values. No evidence for diurnal variation of adult NGF-Rt was apparent. Pregnant women in their third trimester showed significantly elevated urine NGF-Rt values compared with age-matched normals. Affinity labeling of NGF-Rt with 125I-NGF followed by immunoprecipitation with ME20.4-IgG and gel autoradiography indicated that neonatal urine contained high amounts of truncated receptor (Mr = 50 kd); decreasingly lower amounts of NGF-Rt were observed on gel autoradiograms with development, indicating that the two-site radiometric immunosorbent assay correlated well with the affinity labeling technique for measuring NGF-Rt. NGF-Rt in urines from 1-month-old and 36-year-old subjects showed no differences in affinities for NGF or for the monoclonal antibody IIIG5. These data show that NGF-Rt is developmentally regulated in human urine, and are discussed in relation to the development and maturation of the peripheral nervous system.

  6. Nuclear factor-κB mediates placental growth factor induced pro-labour mediators in human placenta.

    PubMed

    Lappas, Martha

    2012-07-01

    Prostaglandins, pro-inflammatory cytokines, extracellular matrix remodelling enzymes and nuclear factor-kappa B (NF-κB) are involved in the mechanisms of term and preterm parturition. Recent studies have reported an increase in angiogenesis-related genes during term and preterm labour, including placental growth factor (PLGF). In non-gestational tissues, PLGF induces inflammation via NF-κB. The aim of this study was to determine the effect of PLGF on the gene expression and release of pro-labour mediators in human placenta. Samples were obtained from normal pregnancies at the time of Caesarean section. Human placenta was incubated in the absence (basal control) or presence of a 10 ng/ml PLGF for 24 h. Inflammatory gene expression was analysed by quantitative RT-PCR, concentration of pro-inflammatory cytokines and prostaglandins was quantified by ELISA, and secretory matrix metalloproteinases (MMPs) activity by zymography. NF-κB DNA-binding activity and IκB-α (inhibitor of NF-κB) protein degradation were analysed by ELISA and Western blotting, respectively. PLGF significantly increased interleukin (IL)-6 and IL-8 gene expression and secretion, cyclooxygenase-2 expression and resultant prostaglandin (PG) E(2) and PGF(2α) release, and MMP-9 gene expression and enzyme production. PLGF induced the degradation of IκB-α whilst increasing NF-κB p65 DNA-binding activity. The PLGF-induced pro-labour responses were abrogated by co-treatment with the NF-κB inhibitor BAY 11-7082. In summary, the pro-inflammatory and pro-labour effects of PLGF in human placenta are mediated by NF-κB.

  7. Mechano-growth factor induces migration of rat mesenchymal stem cells by altering its mechanical properties and activating ERK pathway

    SciTech Connect

    Wu, Jiamin; Wu, Kewen; Lin, Feng; Luo, Qing; Yang, Li; Shi, Yisong; Song, Guanbin; Sung, Kuo-Li Paul

    2013-11-08

    Highlights: •MGF induced the migration of rat MSC in a concentration-dependent manner. •MGF enhanced the mechanical properties of rMSC in inducing its migration. •MGF activated the ERK 1/2 signaling pathway of rMSC in inducing its migration. •rMSC mechanics may synergy with ERK 1/2 pathway in MGF-induced rMSC migration. -- Abstract: Mechano-growth factor (MGF) generated by cells in response to mechanical stimulation has been identified as a mechano effector molecule, playing a key role in regulating mesenchymal stem cell (MSC) function, including proliferation and migration. However, the mechanism(s) underlying how MGF-induced MSC migration occurs is still unclear. In the present study, MGF motivated migration of rat MSCs (rMSCs) in a concentration-dependent manner and optimal concentration of MGF at 50 ng/mL (defined as MGF treatment in this paper) was demonstrated. Notably, enhancement of mechanical properties that is pertinent to cell migration, such as cell traction force and cell stiffness were found to respond to MGF treatment. Furthermore, MGF increased phosphorylation of extracellular signal-regulated kinase (ERK), ERK inhibitor (i.e., PD98059) suppressed ERK phosphorylation, and abolished MGF-induced rMSC migration were found, demonstrating that ERK is involved molecule for MGF-induced rMSC migration. These in vitro evidences of MGF-induced rMSC migration and its direct link to altering rMSC mechanics and activating the ERK pathway, uncover the underlying biomechanical and biological mechanisms of MGF-induced rMSC migration, which may help find MGF-based application of MSC in clinical therapeutics.

  8. The soluble guanylyl cyclase inhibitor NS-2028 reduces vascular endothelial growth factor-induced angiogenesis and permeability.

    PubMed

    Morbidelli, Lucia; Pyriochou, Anastasia; Filippi, Sandra; Vasileiadis, Ioannis; Roussos, Charis; Zhou, Zongmin; Loutrari, Heleni; Waltenberger, Johannes; Stössel, Anne; Giannis, Athanassios; Ziche, Marina; Papapetropoulos, Andreas

    2010-03-01

    Nitric oxide (NO) is known to promote vascular endothelial growth factor (VEGF)-stimulated permeability and angiogenesis. However, effector molecules that operate downstream of NO in this pathway remain poorly characterized. Herein, we determined the effect of soluble guanylyl cyclase (sGC) inhibition on VEGF responses in vitro and in vivo. Treatment of endothelial cells (EC) with VEGF stimulated eNOS phosphorylation and cGMP accumulation; pretreatment with the sGC inhibitor 4H-8-bromo-1,2,4-oxadiazolo(3,4-d)benz(b)(1,4)oxazin-1-one (NS-2028) blunted cGMP levels without affecting VEGF-receptor phosphorylation. Incubation of cells with NS-2028 blocked the mitogenic effects of VEGF. In addition, cells in which sGC was inhibited exhibited no migration and sprouting in response to VEGF. To study the mechanisms through which NS-2028 inhibits EC migration, we determined the effects of alterations in cGMP levels on p38 MAPK. Initially, we observed that inhibition of sGC attenuated VEGF-stimulated activation of p38. In contrast, the addition of 8-Br-cGMP to EC stimulated p38 phosphorylation. The addition of cGMP elevating agents (BAY 41-2272, DETA NO and YC-1) enhanced EC migration. To test whether sGC also mediated the angiogenic effects of VEGF in vivo, we used the rabbit cornea assay. Animals receiving NS-2028 orally displayed a reduced angiogenic response to VEGF. As increased vascular permeability occurs prior to new blood vessel formation, we determined the effect of NS-2028 in vascular leakage. Using a modified Miles assay, we observed that NS-2028 attenuated VEGF-induced permeability. Overall, we provide evidence that sGC mediates the angiogenic and permeability-promoting activities of VEGF, indicating the significance of sGC as a downstream effector of VEGF-triggered responses.

  9. Neuropilin-1 modulates vascular endothelial growth factor-induced poly(ADP-ribose)-polymerase leading to reduced cerebrovascular apoptosis.

    PubMed

    Mey, Lilli; Hörmann, Mareike; Schleicher, Nadine; Reuter, Peter; Dönges, Simone; Kinscherf, Ralf; Gassmann, Max; Gerriets, Tibo; Al-Fakhri, Nadia

    2013-11-01

    Cerebral ischemia is encompassed by cerebrovascular apoptosis, yet the mechanisms behind apoptosis regulation are not fully understood. We previously demonstrated inhibition of endothelial apoptosis by vascular endothelial growth factor (VEGF) through upregulation of poly(ADP-ribose)-polymerase (PARP) expression. However, PARP overactivation through oxidative stress can lead to necrosis. This study tested the hypothesis that neuropilin-1 (NP-1), an alternative VEGF receptor, regulates the response to cerebral ischemia by modulating PARP expression and, in turn, apoptosis inhibition by VEGF. In endothelial cell culture, NP-1 colocalized with VEGF receptor-2 (VEGFR-2) and acted as its coreceptor. This significantly enhanced VEGF-induced PARP mRNA and protein expression demonstrated by receptor-specific inhibitors and VEGF-A isoforms. NP-1 augmented the inhibitory effect of VEGF/VEGFR-2 interaction on apoptosis induced by adhesion inhibition through the αV-integrin inhibitor cRGDfV. NP-1/VEGFR-2 signal transduction involved JNK and Akt. In rat models of permanent and temporary middle cerebral artery occlusion, the ischemic cerebral hemispheres displayed endothelial and neuronal apoptosis next to increased endothelial NP-1 and VEGFR-2 expression compared to non-ischemic cerebral hemispheres, sham-operated or untreated controls. Increased vascular superoxide dismutase-1 and catalase expression as well as decreased glycogen reserves indicated oxidative stress in the ischemic brain. Of note, protein levels of intact PARP remained stable despite pro-apoptotic conditions through increased PARP mRNA production during cerebral ischemia. In conclusion, NP-1 is upregulated in conditions of imminent cerebrovascular apoptosis to reinforce apoptosis inhibition and modulate VEGF-dependent PARP expression and activation. We propose that NP-1 is a key modulator of VEGF maintaining cerebrovascular integrity during ischemia. Modulating the function of NP-1 to target PARP could help to

  10. Controlled microfluidics to examine growth-factor induced migration of neural progenitors in the Drosophila visual system.

    PubMed

    Beck, Cade; Singh, Tanya; Farooqi, Angela; Venkatesh, Tadmiri; Vazquez, Maribel

    2016-03-15

    The developing visual system in Drosophila melanogaster provides an excellent model with which to examine the effects of changing microenvironments on neural cell migration via microfluidics, because the combined experimental system enables direct genetic manipulation, in vivo observation, and in vitro imaging of cells, post-embryo. Exogenous signaling from ligands such as fibroblast growth factor (FGF) is well-known to control glia differentiation, cell migration, and axonal wrapping central to vision. The current study employs a microfluidic device to examine how controlled concentration gradient fields of FGF are able to regulate the migration of vision-critical glia cells with and without cellular contact with neuronal progenitors. Our findings quantitatively illustrate a concentration-gradient dependent chemotaxis toward FGF, and further demonstrate that glia require collective and coordinated neuronal locomotion to achieve directionality, sustain motility, and propagate long cell distances in the visual system. Conventional assays are unable to examine concentration- and gradient-dependent migration. Our data illustrate quantitative correlations between ligand concentration/gradient and glial cell distance traveled, independent or in contact with neurons. Microfluidic systems in combination with a genetically-amenable experimental system empowers researchers to dissect the signaling pathways that underlie cellular migration during nervous system development. Our findings illustrate the need for coordinated neuron-glia migration in the Drosophila visual system, as only glia within heterogeneous populations exhibited increasing motility along distances that increased with increasing FGF concentration. Such coordinated migration and chemotactic dependence can be manipulated for potential therapeutic avenues for NS repair and/or disease treatment. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Hepatocyte growth factor/scatter factor induces a variety of tissue- specific morphogenic programs in epithelial cells

    PubMed Central

    1995-01-01

    Hepatocyte growth factor/scatter factor (HGF/SF) is the mesenchymal ligand of the epithelial tyrosine kinase receptor c-Met. In vitro, HGF/SF has morphogenic properties, e.g., induces kidney epithelial cells to form branching ducts in collagen gels. Mutation of the HGF/SF gene in mice results in embryonic lethality due to severe liver and placenta defects. Here, we have evaluated the morphogenic activity of HGF/SF with a large variety of epithelial cells grown in three- dimensional collagen matrices. We found that HGF/SF induces SW 1222 colon carcinoma cells to form crypt-like structures. In these organoids, cells exhibit apical/basolateral polarity and build a well- developed brush border towards the lumen. Capan 2 pancreas carcinoma cells, upon addition of HGF/SF, develop large hollow spheroids lined with a tight layer of polarized cells. Collagen inside the cysts is digested and the cells show features of pancreatic ducts. HGF/SF induces EpH4 mammary epithelial cells to form long branches with end- buds that resemble developing mammary ducts. pRNS-1-1 prostate epithelial cells in the presence of HGF/SF develop long ducts with distal branching as found in the prostate. Finally, HGF/SF simulates alveolar differentiation in LX-1 lung carcinoma cells. Expression of transfected HGF/SF cDNA in LX-1 lung carcinoma and EpH4 mammary epithelial cells induce morphogenesis in an autocrine manner. In the cell lines tested, HGF/SF activated the Met receptor by phosphorylation of tyrosine residues. These data show that HGF/SF induces intrinsic, tissue-specific morphogenic activities in a wide variety of epithelial cells. Apparently, HGF/SF triggers respective endogenous programs and is thus an inductive, not an instructive, mesenchymal effector for epithelial morphogenesis. PMID:8522613

  12. Nonsteroidal anti-inflammatory drugs attenuate proliferation of colonic carcinoma cells by blocking epidermal growth factor-induced Ca++ mobilization.

    PubMed

    Kokoska, E R; Smith, G S; Miller, T A

    2000-01-01

    Numerous studies suggest that nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit colorectal carcinogenesis. We have previously reported that NSAIDs, in human colonic carcinoma cells (Caco-2), attenuate epidermal growth factor (EGF)-induced cellular proliferation through a process independent of their inhibitory effects on prostaglandin synthesis. Furthermore, separate studies have also suggested that NSAIDs inhibit EGF-induced store-operated Ca++ influx. Thus we developed the hypothesis that NSAIDs may limit the activity of EGF by altering intracellular Ca++ ([Ca++]i) mobilization. Serum-deprived Caco-2 cells were employed for all experimentation. [Ca++]i was measured with Fluo-3 and extracellular Ca++ influx was monitored by quenching Fluo-3 fluorescence with Mn++. Proliferation was quantitated with two assays: cellular nucleic acid and total protein content. Caco-2 cells exposed to EGF demonstrated an initial increase in [Ca++]i which was blocked by neomycin, an inhibitor of IPsubscript 3 generation, and the phospholipase C inhibitor U73122 but not U73343 (inactive control). This was followed by sustained extracellular Ca++ influx, which was attenuated with calcium-free buffer (-Ca++), the store- operated Ca++ channel blocker lanthanum, indomethacin, ibuprofen, and aspirin. In subsequent studies, cells were treated with either serum-free media or EGF +/- the aforementioned inhibitors, and again serum starved. Cells exposed to EGF +/- the inactive phospholipase C inhibitor U73343 demonstrated a significant increase in nucleic acid and protein. However, proliferation induced by EGF was not observed when [Ca++]i elevation was prevented by blocking either internal Ca++ store release via phospholipase C/IPsubscript 3 or sustained Ca++ influx through store-operated Ca++ channels. Sustained [Ca++]i elevation, as induced by EGF, appears to be required for mitogenesis. These data support our premise that one mechanism whereby NSAIDs may attenuate colonic neoplasia is

  13. Synergistic effects of methylnaltrexone with 5-fluorouracil and bevacizumab on inhibition of vascular endothelial growth factor-induced angiogenesis.

    PubMed

    Singleton, Patrick A; Garcia, Joe G N; Moss, Jonathan

    2008-06-01

    Many patients with cancer receive combinations of drug treatments that include 5-fluorouracil (5-FU) and bevacizumab. Therapeutic doses of 5-FU are often associated with unwanted side effects, and bevacizumab is costly. Therefore, we explored potential agents that can reduce the therapeutic concentration of these drugs. Our data indicate that methylnaltrexone (MNTX), a peripheral antagonist of the mu-opioid receptor, exerts a synergistic effect with 5-FU and bevacizumab on inhibition of vascular endothelial growth factor (VEGF)-induced human pulmonary microvascular endothelial cell (EC) proliferation and migration, two key components in cancer-associated angiogenesis. MNTX inhibited EC proliferation with an IC(50) of approximately 100 nmol/L. Adding 100 nmol/L MNTX to EC shifted the IC(50) of 5-FU from approximately 5 micromol/L to approximately 7 nmol/L. Further, adding 50 ng/mL MNTX shifted the IC(50) of bevacizumab on inhibition of EC migration from approximately 25 to approximately 6 ng/mL. These synergistic effects were not observed with naltrexone, a tertiary mu-opioid receptor antagonist. On a mechanistic level, we observed that treatment of human EC with MNTX, but not naltrexone, increased receptor protein tyrosine phosphatase mu activity, which was independent of mu-opioid receptor expression. Silencing receptor protein tyrosine phosphatase mu expression (small interfering RNA) in human EC inhibited both synergy between MNTX and bevacizumab or 5-FU and increased VEGF-induced tyrosine phosphorylation of Src and p190 RhoGAP with enhanced activation of Akt and the actin cytoskeletal regulatory protein, RhoA, whereas silencing Src, Akt, or RhoA blocked VEGF-induced angiogenic events. Therefore, addition of MNTX could potentially lower the therapeutic doses of 5-FU and bevacizumab, which could improve index.

  14. Increased axonal regeneration through a biodegradable amnionic tube nerve conduit: effect of local delivery and incorporation of nerve growth factor/hyaluronic acid media.

    PubMed

    Mohammad, J A; Warnke, P H; Pan, Y C; Shenaq, S

    2000-01-01

    The authors emphasize the possible pharmacological enhancement of axonal regeneration using a specific growth factor/ extracellular media incorporated in a biodegradable nonneural nerve conduit material. They investigated the early effects on nerve regeneration of continuous local delivery of nerve growth factor (NGF) and the local incorporation of hyaluronic acid (HA) inside a newly manufactured nerve conduit material from fresh human amnionic membrane. Human amnionic membrane contains important biochemical factors that play a major neurotrophic role in the nerve regeneration process. The process of manufacturing a nerve conduit from fresh human amnionic membrane is described. This nerve conduit system was used in rabbits to bridge a 25-mm nerve gap over 3 months. NGF was released locally, over 28 days, at the distal end of the tube via a system of slow release, and HA was incorporated inside the lumen of the tube at the time of surgery. NGF/HA treatment promoted axonal regeneration across the amnionic tube nerve conduit (8,962 +/- 383 myelinated axons) 45% better than the nontreated amnionic tube group (6,180 +/- 353 myelinated axons). The authors demonstrate that NGF/HA media enhances additional axonal regeneration in the amnionic tube nerve conduit. This result is secondary to the effect of the amnion promoting biochemical factors, in combination with the NGF/HA effect on facilitating early events in the nerve regeneration process.

  15. The chemokine CXCL12 mediates the anti-amyloidogenic action of painless human nerve growth factor

    PubMed Central

    Capsoni, Simona; Malerba, Francesca; Carucci, Nicola Maria; Rizzi, Caterina; Criscuolo, Chiara; Origlia, Nicola; Calvello, Mariantonietta; Viegi, Alessandro; Meli, Giovanni

    2017-01-01

    Abstract Nerve growth factor is a therapeutic candidate for Alzheimer’s disease. Due to its pain-inducing activity, in current clinical trials nerve growth factor is delivered locally into the brain by neurosurgery, but data on the efficacy of local nerve growth factor delivery in decreasing amyloid-β deposition are not available. To reduce the nerve growth factor pain-inducing side effects, thus avoiding the need for local brain injection, we developed human painless nerve growth factor (hNGFp), inspired by the human genetic disease hereditary sensory and autonomic neuropathy type V. hNGFp has identical neurotrophic potency as wild-type human nerve growth factor, but a 10-fold lower pain sensitizing activity. In this study we first mimicked, in the 5xFAD mouse model, the intraparenchymal delivery of hNGFp used in clinical trials and found it to be ineffective in decreasing amyloid-β plaque load. On the contrary, the same dose of hNGFp delivered intranasally, which was widely biodistributed in the brain and did not induce pain, showed a potent anti-amyloidogenic action and rescued synaptic plasticity and memory deficits. We found that hNGFp acts on glial cells, modulating inflammatory proteins such as the soluble TNFα receptor II and the chemokine CXCL12. We further established that the rescuing effect by hNGFp is mediated by CXCL12, as pharmacological inhibition of CXCL12 receptor CXCR4 occludes most of hNGFp effects. These findings have significant therapeutic implications: (i) we established that a widespread exposure of the brain is required for nerve growth factor to fully exert its neuroprotective actions; and (ii) we have identified a new anti-neurodegenerative pathway as a broad target for new therapeutic opportunities for neurodegenerative diseases. PMID:28031222

  16. The chemokine CXCL12 mediates the anti-amyloidogenic action of painless human nerve growth factor.

    PubMed

    Capsoni, Simona; Malerba, Francesca; Carucci, Nicola Maria; Rizzi, Caterina; Criscuolo, Chiara; Origlia, Nicola; Calvello, Mariantonietta; Viegi, Alessandro; Meli, Giovanni; Cattaneo, Antonino

    2017-01-01

    Nerve growth factor is a therapeutic candidate for Alzheimer's disease. Due to its pain-inducing activity, in current clinical trials nerve growth factor is delivered locally into the brain by neurosurgery, but data on the efficacy of local nerve growth factor delivery in decreasing amyloid-β deposition are not available. To reduce the nerve growth factor pain-inducing side effects, thus avoiding the need for local brain injection, we developed human painless nerve growth factor (hNGFp), inspired by the human genetic disease hereditary sensory and autonomic neuropathy type V. hNGFp has identical neurotrophic potency as wild-type human nerve growth factor, but a 10-fold lower pain sensitizing activity. In this study we first mimicked, in the 5xFAD mouse model, the intraparenchymal delivery of hNGFp used in clinical trials and found it to be ineffective in decreasing amyloid-β plaque load. On the contrary, the same dose of hNGFp delivered intranasally, which was widely biodistributed in the brain and did not induce pain, showed a potent anti-amyloidogenic action and rescued synaptic plasticity and memory deficits. We found that hNGFp acts on glial cells, modulating inflammatory proteins such as the soluble TNFα receptor II and the chemokine CXCL12. We further established that the rescuing effect by hNGFp is mediated by CXCL12, as pharmacological inhibition of CXCL12 receptor CXCR4 occludes most of hNGFp effects. These findings have significant therapeutic implications: (i) we established that a widespread exposure of the brain is required for nerve growth factor to fully exert its neuroprotective actions; and (ii) we have identified a new anti-neurodegenerative pathway as a broad target for new therapeutic opportunities for neurodegenerative diseases.

  17. Intranasal nerve growth factor bypasses the blood-brain barrier and affects spinal cord neurons in spinal cord injury

    PubMed Central

    Aloe, Luigi; Bianchi, Patrizia; De Bellis, Alberto; Soligo, Marzia; Rocco, Maria Luisa

    2014-01-01

    The purpose of this work was to investigate whether, by intranasal administration, the nerve growth factor bypasses the blood-brain barrier and turns over the spinal cord neurons and if such therapeutic approach could be of value in the treatment of spinal cord injury. Adult Sprague-Dawley rats with intact and injured spinal cord received daily intranasal nerve growth factor administration in both nostrils for 1 day or for 3 consecutive weeks. We found an increased content of nerve growth factor and enhanced expression of nerve growth factor receptor in the spinal cord 24 hours after a single intranasal administration of nerve growth factor in healthy rats, while daily treatment for 3 weeks in a model of spinal cord injury improved the deficits in locomotor behaviour and increased spinal content of both nerve growth factor and nerve growth factor receptors. These outcomes suggest that the intranasal nerve growth factor bypasses blood-brain barrier and affects spinal cord neurons in spinal cord injury. They also suggest exploiting the possible therapeutic role of intranasally delivered nerve growth factor for the neuroprotection of damaged spinal nerve cells. PMID:25206755

  18. Integrin antagonists affect growth and pathfinding of ventral motor nerves in the trunk of embryonic zebrafish.

    PubMed

    Becker, Thomas; McLane, Mary Ann; Becker, Catherina G

    2003-05-01

    Integrins are thought to be important receptors for extracellular matrix (ECM) components on growing axons. Ventral motor axons in the trunk of embryonic zebrafish grow in a midsegmental pathway through an environment rich in ECM components. To test the role of integrins in this process, integrin antagonists (the disintegrin echistatin in native and recombinant form, as well as the Arg-Gly-Asp-Ser peptide) were injected into embryos just prior to axon outgrowth at 14-16 h postfertilization (hpf). All integrin antagonists affected growth of ventral motor nerves in a similar way and native echistatin was most effective. At 24 hpf, when only the three primary motor axons per trunk hemisegment had grown out, 80% (16 of 20) of the embryos analyzed had abnormal motor nerves after injection of native echistatin, corresponding to 19% (91 of 480) of all nerves. At 33 hpf, when secondary motor axons were present in the pathway, 100% of the embryos were affected (24 of 24), with 20% of all nerves analyzed (196 of 960) being abnormal. Phenotypes comprised abnormal branching (64% of all abnormal nerves) and truncations (36% of all abnormal nerves) of ventral motor nerves at 24 hpf and mostly branching of the nerves at 33 hpf (94% of all abnormal nerves). Caudal branches were at least twice as frequent as rostral branches. Surrounding trunk tissue and a number of other axon fascicles were apparently not affected by the injections. Thus integrin function contributes to both growth and pathfinding of axons in ventral motor nerves in the trunk of zebrafish in vivo.

  19. Nerve growth factor facilitates redistribution of adrenergic and non-adrenergic non-cholinergic perivascular nerves injured by phenol in rat mesenteric resistance arteries.

    PubMed

    Yokomizo, Ayako; Takatori, Shingo; Hashikawa-Hobara, Narumi; Goda, Mitsuhiro; Kawasaki, Hiromu

    2016-01-05

    We previously reported that nerve growth factor (NGF) facilitated perivascular sympathetic neuropeptide Y (NPY)- and calcitonin gene-related peptide (CGRP)-containing nerves injured by the topical application of phenol in the rat mesenteric artery. We also demonstrated that mesenteric arterial nerves were distributed into tyrosine hydroxylase (TH)-, substance P (SP)-, and neuronal nitric oxide synthase (nNOS)-containing nerves, which had axo-axonal interactions. In the present study, we examined the effects of NGF on phenol-injured perivascular nerves, including TH-, NPY-, nNOS-, CGRP-, and SP-containing nerves, in rat mesenteric arteries in more detail. Wistar rats underwent the in vivo topical application of 10% phenol to the superior mesenteric artery, proximal to the abdominal aorta, under pentobarbital-Na anesthesia. The distribution of perivascular nerves in the mesenteric arteries of the 2nd to 3rd-order branches isolated from 8-week-old Wistar rats was investigated immunohistochemically using antibodies against TH-, NPY-, nNOS-, CGRP-, and SP-containing nerves. The topical phenol treatment markedly reduced the density of all nerves in these arteries. The administration of NGF at a dose of 20µg/kg/day with an osmotic pump for 7 days significantly increased the density of all perivascular nerves over that of sham control levels. These results suggest that NGF facilitates the reinnervation of all perivascular nerves injured by phenol in small resistance arteries.

  20. Nerve Growth Factor Inhibits Sympathetic Neurons' Response to an Injury Cytokine

    NASA Astrophysics Data System (ADS)

    Shadiack, Annette M.; Vaccariello, Stacey A.; Sun, Yi; Zigmond, Richard E.

    1998-06-01

    Axonal damage to adult peripheral neurons causes changes in neuronal gene expression. For example, axotomized sympathetic, sensory, and motor neurons begin to express galanin mRNA and protein, and recent evidence suggests that galanin plays a role in peripheral nerve regeneration. Previous studies in sympathetic and sensory neurons have established that galanin expression is triggered by two consequences of nerve transection: the induction of leukemia inhibitory factor (LIF) and the reduction in the availability of the target-derived factor, nerve growth factor. It is shown in the present study that no stimulation of galanin expression occurs following direct application of LIF to intact neurons in the superior cervical sympathetic ganglion. Injection of animals with an antiserum to nerve growth factor concomitant with the application of LIF, on the other hand, does stimulate galanin expression. The data suggest that the response of neurons to an injury factor, LIF, is affected by whether the neurons still receive trophic signals from their targets.

  1. Nerve growth factor from cobra venom inhibits the growth of Ehrlich tumor in mice.

    PubMed

    Osipov, Alexey V; Terpinskaya, Tatiana I; Kryukova, Elena V; Ulaschik, Vladimir S; Paulovets, Lubov V; Petrova, Elena A; Blagun, Ekaterina V; Starkov, Vladislav G; Utkin, Yuri N

    2014-02-26

    The effects of nerve growth factor (NGF) from cobra venom (cvNGF) on growth of Ehrlich ascites carcinoma (EAC) cells inoculated subcutaneously in mice have been studied. The carcinoma growth slows down, but does not stop, during a course of cvNGF injections and restores after the course has been discontinued. The maximal anti-tumor effect has been observed at a dose of 8 nmoles cvNGF/kg body weight. cvNGF does not impact on lifespan of mice with grafted EAC cells. K252a, a tyrosine kinase inhibitor, attenuates the anti-tumor effect of cvNGF indicating the involvement of TrkA receptors in the process. cvNGF has induced also increase in body weight of the experimental animals. In overall, cvNGF shows the anti-tumor and weight-increasing effects which are opposite to those described for mammalian NGF (mNGF). However in experiments on breast cancer cell line MCF-7 cvNGF showed the same proliferative effects as mNGF and had no cytotoxic action on tumor cells in vitro. These data suggest that cvNGF slows down EAC growth via an indirect mechanism in which TrkA receptors are involved.

  2. Nerve Growth Factor from Cobra Venom Inhibits the Growth of Ehrlich Tumor in Mice

    PubMed Central

    Osipov, Alexey V.; Terpinskaya, Tatiana I.; Kryukova, Elena V.; Ulaschik, Vladimir S.; Paulovets, Lubov V.; Petrova, Elena A.; Blagun, Ekaterina V.; Starkov, Vladislav G.; Utkin, Yuri N.

    2014-01-01

    The effects of nerve growth factor (NGF) from cobra venom (cvNGF) on growth of Ehrlich ascites carcinoma (EAC) cells inoculated subcutaneously in mice have been studied. The carcinoma growth slows down, but does not stop, during a course of cvNGF injections and restores after the course has been discontinued. The maximal anti-tumor effect has been observed at a dose of 8 nmoles cvNGF/kg body weight. cvNGF does not impact on lifespan of mice with grafted EAC cells. K252a, a tyrosine kinase inhibitor, attenuates the anti-tumor effect of cvNGF indicating the involvement of TrkA receptors in the process. cvNGF has induced also increase in body weight of the experimental animals. In overall, cvNGF shows the anti-tumor and weight-increasing effects which are opposite to those described for mammalian NGF (mNGF). However in experiments on breast cancer cell line MCF-7 cvNGF showed the same proliferative effects as mNGF and had no cytotoxic action on tumor cells in vitro. These data suggest that cvNGF slows down EAC growth via an indirect mechanism in which TrkA receptors are involved. PMID:24577582

  3. Role of CD44 in Malignant Peripheral Nerve Sheath Tumor Growth and Metastasis

    DTIC Science & Technology

    2002-09-01

    Malignant peripheral nerve sheath tumors ( MPNSTs ) are aggressive malignancies that arise within peripheral nerves. These tumors occur with increased...and abnormal expression of the epidermal growth factor receptor (EGFR). We previously found that MPNSTs express increased levels of the CD44 family...kinase activity (and not increased Ras-GTP) contributes to MPNST cell invasion. We further find that EGFR contributes at least part of the elevated Src

  4. Neurotrophins enhance electric field-directed growth cone guidance and directed nerve branching.

    PubMed

    McCaig, C D; Sangster, L; Stewart, R

    2000-03-01

    Neurotrophins play major roles in the developing nervous system in controlling neuronal differentiation, neurite outgrowth, guidance and branching, synapse formation and maturation, and neuronal survival or death. There is increasing evidence that nervous system construction takes place in the presence of dc electric fields, which fluctuate dynamically in space and time during embryonic development. These have their origins in the neural tube itself, as well as in surrounding skin and gut. Early disruption of these endogenous electric fields causes failure of the nervous system to form, or else it forms aberrantly. Nerve growth, guidance, and branching are controlled tightly during pathway construction and in vitro dc electric fields have profound effects on each of these behaviours. We have used cultured neurones to ask whether neurotrophins and dc electric fields might interact in shaping neuronal growth, given their coexistence in vivo. Electric field-directed nerve growth generally was enhanced by the simultaneous presentation of several neurotrophins to the growth cone. Under certain circumstances, more nerves turned cathodally, they turned faster, further, and in lower field strengths. Intriguingly, other combinations of dc electric field and neurotrophins (low field strength and neurotrophin 3 (NT-3) switched the direction of growth cone turning. Additionally, cathodally directed nerve growth was faster and directed branching was much more common when electric fields and neurotrophins interacted with neuronal growth cones. Given such profound changes in growth cone behaviour in vitro, neurotrophins and endogenous electric fields are likely to interact in vivo.

  5. Nerve growth factor promotes in vitro proliferation of neural stem cells from tree shrews

    PubMed Central

    Xiong, Liu-lin; Chen, Zhi-wei; Wang, Ting-hua

    2016-01-01

    Neural stem cells promote neuronal regeneration and repair of brain tissue after injury, but have limited resources and proliferative ability in vivo. We hypothesized that nerve growth factor would promote in vitro proliferation of neural stem cells derived from the tree shrews, a primate-like mammal that has been proposed as an alternative to primates in biomedical translational research. We cultured neural stem cells from the hippocampus of tree shrews at embryonic day 38, and added nerve growth factor (100 μg/L) to the culture medium. Neural stem cells from the hippocampus of tree shrews cultured without nerve growth factor were used as controls. After 3 days, fluorescence microscopy after DAPI and nestin staining revealed that the number of neurospheres and DAPI/nestin-positive cells was markedly greater in the nerve growth factor-treated cells than in control cells. These findings demonstrate that nerve growth factor promotes the proliferation of neural stem cells derived from tree shrews. PMID:27212919

  6. Sustained Growth Factor Delivery Promotes Axonal Regeneration in Long Gap Peripheral Nerve Repair

    PubMed Central

    Kokai, Lauren E.; Bourbeau, Dennis; Weber, Douglas; McAtee, Jedidiah

    2011-01-01

    The aim of this study was to evaluate the long-term effect of localized growth factor delivery on sciatic nerve regeneration in a critical-size (>1 cm) peripheral nerve defect. Previous work has demonstrated that bioactive proteins can be encapsulated within double-walled, poly(lactic-co-glycolic acid)/poly(lactide) microspheres and embedded within walls of biodegradable polymer nerve guides composed of poly(caprolactone). Within this study, nerve guides containing glial cell line-derived neurotrophic factor (GDNF) were used to bridge a 1.5-cm defect in the male Lewis rat for a 16-week period. Nerve repair was evaluated through functional assessment of joint angle range of motion using video gait kinematics, gastrocnemius twitch force, and gastrocnemius wet weight. Histological evaluation of nerve repair included assessment of Schwann cell and neurofilament location with immunohistochemistry, evaluation of tissue integration and organization throughout the lumen of the regenerated nerve with Masson's trichrome stain, and quantification of axon fiber density and g-ratio. Results from this study showed that the measured gastrocnemius twitch force in animals treated with GDNF was significantly higher than negative controls and was not significantly different from the isograft-positive control group. Histological assessment of explanted conduits after 16 weeks showed improved tissue integration within GDNF releasing nerve guides compared to negative controls. Nerve fibers were present across the entire length of GDNF releasing guides, whereas nerve fibers were not detectable beyond the middle region of negative control guides. Therefore, our results support the use of GDNF for improved functional recovery above negative controls following large axonal defects in the peripheral nervous system. PMID:21189072

  7. Effects of nerve growth factor delivery via a gel to inferior alveolar nerve in mandibular distraction osteogenesis.

    PubMed

    Wang, Lei; Cao, Jian; Lei, De-lin; Cheng, Xiao-bing; Yang, Yao-wu; Hou, Rui; Zhao, Ying-hua; Cui, Fu-zhai

    2009-11-01

    Inferior alveolar nerve (IAN) injury is a concern in mandible distraction osteogenesis (DO). We have previously demonstrated that repeated local injections of human nerve growth factor beta (NGF-beta) have significantly enhanced the histologic recovery of the IAN in a rabbit model of DO. This study was to further test the effect of a single injection of human NGF-beta delivered via a collagen/nanohydroxyapatite/kappa-carrageenan gel to the recovery of the IAN in DO. Rabbits underwent mandibular DO at a rate of 0.75 mm/12 h for 6 days. At the end of the distraction period, injections were performed near the IAN percutaneously as follows: group 1, human NGF-beta in the gel; group 2, human NGF-beta in saline; group 3, the gel alone; and group 4, saline alone. At 14 days after the end of distraction, IAN histologic findings and histomorphometric parameters were evaluated. Histologically, there were less myelin debris and more abundant regenerating nerve fibers in group 1 than the other groups. Both the myelinated fiber density and the myelinated axon area in group 1 were significantly higher than groups 3 and 4 (P < 0.01); the myelinated axon area in the group 1 was significantly higher than group 2 (P < 0.01). In conclusion, the delivery of human NGF-beta in the gel leads to a better acceleration of the IAN injury recovery over the saline delivery. It provides a possible way to enhance the recovery of nerve injuries in craniofacial DO clinically.

  8. Tumor necrosis factor-like weak inducer of apoptosis and its receptor fibroblast growth factor-inducible 14 are expressed in urticarial vasculitis.

    PubMed

    Li, Mengmeng; Chen, Tao; Guo, Zaipei; Li, Jingyi; Cao, Na

    2013-11-01

    Tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK), a member of the TNF family, has been implicated as a pro-inflammatory cytokine in many types of autoimmune and infectious diseases. However, information about TWEAK in dermatological diseases is limited. To date, no studies have investigated the roles of TWEAK in patients with urticarial vasculitis (UV). This study aimed to assess serum TWEAK levels, together with TWEAK and fibroblast growth factor-inducible 14 (Fn14) expressions of skin lesions in patients with UV. Serum TWEAK levels in patients with UV, together with patients with cutaneous leukocytoclastic angiitis (CLA) and healthy controls were detected by enzyme-linked immunosorbent assay; TWEAK and Fn14 expressions of skin lesions were analyzed by immunohistochemistry. Results showed that TWEAK and Fn14 were abundantly expressed in the dermal vessel wall of lesional skin in patients with UV but not healthy controls. Serum TWEAK levels in the acute stage in patients with UV were significantly higher than those in the convalescent stage and healthy controls. Serum TWEAK levels were elevated significantly in patients with CLA compared with those in healthy controls. Our previous study indicated that TWEAK may be an important mediator for the development of vascular inflammation in skin. In addition, we also found that TWEAK blockade substantially reduced vascular damage and perivascular leukocyte infiltrates in lipopolysaccharide-induced cutaneous vasculitis. Our study shows that TWEAK may be associated with the pathogenesis of UV; it is therefore suggested that TWEAK may be a potential therapeutic target for UV and other types of cutaneous vasculitis. © 2013 Japanese Dermatological Association.

  9. ATF3 increases the intrinsic growth state of DRG neurons to enhance peripheral nerve regeneration.

    PubMed

    Seijffers, Rhona; Mills, Charles D; Woolf, Clifford J

    2007-07-25

    Peripheral axons of dorsal root ganglion (DRG) neurons, but not their central axons in the dorsal columns, regenerate after injury. However, if the neurons are conditioned by a peripheral nerve injury into an actively growing state, the rate of peripheral axonal growth is accelerated and the injured central axons begin to regenerate. The growth-promoting effects of conditioning injuries have two components, increased axonal growth and a reduced response to inhibitory myelin cues. We have examined which transcription factors activated by peripheral axonal injury may mediate the conditioning effect by regulating expression of effectors that increase the intrinsic growth state of the neurons. Activating transcription factor 3 (ATF3) is a prime candidate because it is induced in all injured DRG neurons after peripheral, but not central, axonal damage. To investigate if ATF3 promotes regeneration, we generated transgenic mice that constitutively express this transcription factor in non-injured adult DRG neurons. The rate of peripheral nerve regeneration was enhanced in the transgenic mice to an extent comparable to that produced by a preconditioning nerve injury. The expression of some growth-associated genes, such as SPRR1A, but not others like GAP-43, was increased in the non-injured neurons. ATF3 increased DRG neurite elongation when cultured on permissive substrates but did not overcome the inhibitory effects of myelin or promote central axonal regeneration in the spinal cord in vivo. We conclude that ATF3 contributes to nerve regeneration by increasing the intrinsic growth state of injured neurons.

  10. Transforming Growth Factor-β Promotes Axonal Regeneration After Chronic Nerve Injury.

    PubMed

    Sulaiman, Wale A R

    2016-04-01

    When spinal cord injury (SCI) occurs, injured cells must survive and regenerate to close gaps caused by the injury and to create functional motor units. After peripheral nerve injury, Wallerian degeneration in the distal nerve stump creates a neurotrophic and growth-supportive environment for injured neurons and axons via Schwann cells and secreted cytokines/neurotrophins. In both SCI and peripheral nerve injury, injured motor and sensory neurons must regenerate axons, eventually reaching and reinnervating target tissue (SDC Figure 1, http://links.lww.com/BRS/B116). This process is often unsuccessful after SCI, and the highly complex anatomy of branching axons and nerves in the peripheral nervous system leads to slow recovery of function, even with careful and appropriate techniques.

  11. Increased vascular penetration and nerve growth in the meniscus: a potential source of pain in osteoarthritis.

    PubMed

    Ashraf, Sadaf; Wibberley, Helen; Mapp, Paul Ian; Hill, Roger; Wilson, Deborah; Walsh, David Andrew

    2011-03-01

    Meniscal damage is a recognised feature of knee osteoarthritis (OA), although its clinical relevance remains uncertain. This study describes vascular penetration and nerve growth in human menisci, providing a potential mechanism for the genesis of pain in knee OA. Menisci obtained post mortem were screened on the basis of high or low macroscopic tibiofemoral chondropathy as a measure of the presence and degree of OA. Forty cases (20 per group) were selected for the study of meniscal vascularity, and 16 (eight per group) for the study of meniscal innervation. Antibodies directed against α-actin and calcitonin gene-related peptide (CGRP) were used to localise blood vessels and nerves by histochemistry. Image analysis was used to compare vascular and nerve densities between groups. Data are presented as median (IQR). Menisci from knees with high chondropathy displayed degeneration of collagen bundles in their outer regions, which were more vascular than the inner regions, with an abrupt decrease in vascularity at the fibrocartilage junction. Vascular densities were increased in menisci from the high compared with low chondropathy group both in the synovium (3.8% (IQR 2.6-5.2), 2.0% (IQR 1.4-2.9), p=0.002) and at the fibrocartilage junction (2.3% (IQR 1.7-3.1), 1.1% (IQR 0.8-1.9), p=0.003), with a greater density of perivascular sensory nerve profiles in the outer region (high chondropathy group, 144 nerve profiles/mm(2) (IQR 134-189); low chondropathy group, 119 nerve profiles/mm(2) (IQR 104-144), p=0.049). Tibiofemoral chondropathy is associated with altered matrix structure, increased vascular penetration, and increased sensory nerve densities in the medial meniscus. The authors suggest therefore that angiogenesis and associated sensory nerve growth in menisci may contribute to pain in knee OA.

  12. Nerve growth factor-mediated vascular endothelial growth factor expression of astrocyte in retinal vascular development.

    PubMed

    Kim, You Sun; Jo, Dong Hyun; Lee, Hanjae; Kim, Jin Hyoung; Kim, Kyu-Won; Kim, Jeong Hun

    2013-02-22

    The angiogenic aspect of neurotrophins and their receptors rather than the neuroscientific aspect has been focused. However, their role in retinal vascular development is underdiscovered. The purpose of this study is to understand the role of neurotrophin receptors in retinal vascular development and the mechanisms of their action. To identify the expression of tropomyosin receptor kinase receptor (Trk) in developing retina, tissues of 4, 8, 12, 16 and 26 day-old mice were prepared for experiments. Immunohistochemistry and immunofluorescence double staining against glial fibrillary acidic protein and type IV collagen were performed. TrkA was expressed mainly along the vessel structure in inner part of retina, especially in retinal astrocyte. In cultured primary astrocyte, recombinant nerve growth factor (NGF) was used to activate TrkA. NGF induced the phosphorylation of TrkA, and it also enhanced the level of activated Akt and vascular endothelial growth factor (VEGF) mRNA. Inhibition of phosphoinositide 3-kinase (PI3K) reversed the NGF-induced activation of these two molecules. This study demonstrated that TrkA activation on NGF leads to VEGF elevation by PI3K-Akt pathway and therefore suggested that TrkA could be a stimulator of retinal vascular development. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. Silk fibroin matrices for the controlled release of nerve growth factor (NGF).

    PubMed

    Uebersax, Lorenz; Mattotti, Marta; Papaloïzos, Michaël; Merkle, Hans P; Gander, Bruno; Meinel, Lorenz

    2007-10-01

    Nerve conduits (NC) for peripheral nerve repair should guide the sprouting axons and physically protect the axonal cone from any damage. The NC should also degrade after completion of its function to obviate the need of subsequent explanation and should optionally be suitable for controlled drug release of embedded growth factors to enhance nerve regeneration. Silk fibroin (SF) is a biocompatible and slowly biodegradable biomaterial with excellent mechanical properties that could meet the above stated requirements. SF material (films) supported the adherence and metabolic activity of PC12 cells, and, in combination with nerve growth factor (NGF), supported neurite outgrowth during PC12 cell differentiation. NGF-loaded SF-NC were prepared from aqueous solutions of NGF and SF (20%, w/w), which were air-dried or freeze-dried (freezing at -20 or -196 degrees C) in suitable molds. NGF release from the three differently prepared SF-NC was prolonged over at least 3 weeks, but the total amount released depended on the drying procedure of the NC. The potency of released NGF was retained within all formulations. Control experiments with differently dried NGF-lactose solutions did not evidence marked protein aggregation (SEC, HPLC), loss of ELISA-reactivity or PC12 cell bioactivity. This study encourages the further exploitation of SF-NC for growth factor delivery and evaluation in peripheral nerve repair.

  14. Growth of injured rabbit optic axons within their degenerating optic nerve

    SciTech Connect

    Lavie, V.; Murray, M.; Solomon, A.; Ben-Bassat, S.; Belkin, M.; Rumelt, S.; Schwartz, M. )

    1990-08-15

    Spontaneous growth of axons after injury is extremely limited in the mammalian central nervous system (CNS). It is now clear, however, that injured CNS axons can be induced to elongate when provided with a suitable environment. Thus injured CNS axons can elongate, but they do not do so unless their environment is altered. We now show apparent regenerative growth of injured optic axons. This growth is achieved in the adult rabbit optic nerve by the use of a combined treatment consisting of: (1) supplying soluble substances originating from growing axons to be injured rabbit optic nerves, and (2) application of low energy He-Ne laser irradiation, which appears to delay degenerative changes in the injured axons. Two to 8 weeks after this treatment, unmyelinated and thinly myelinated axons are found at the lesion site and distal to it. Morphological and immunocytochemical evidence indicate that these thinly myelinated and unmyelinated axons are growing in close association with glial cells. Only these axons are identified as being growing axons. These newly growing axons transverse the site of injury and extend into the distal stump of the nerve, which contains degenerating axons. Axons of this type could be detected distal to the lesion only in nerves subjected to the combined treatment. No unmyelinated or thinly myelinated axons in association with glial cells were seen at 6 or 8 weeks postoperatively in nerves that were not treated, or in nerves in which the two stumps were completely disconnected. Two millimeters distal to the site of injury, the growing axons are confined to a compartment comprising 5%-30% of the cross section of the nerve. A temporal analysis indicates that axons have grown as far as 6 mm distal to the site of injury, by 8 weeks postoperatively.

  15. Magnetic Nanoparticles for Efficient Delivery of Growth Factors: Stimulation of Peripheral Nerve Regeneration.

    PubMed

    Giannaccini, Martina; Calatayud, M Pilar; Poggetti, Andrea; Corbianco, Silvia; Novelli, Michela; Paoli, Melania; Battistini, Pietro; Castagna, Maura; Dente, Luciana; Parchi, Paolo; Lisanti, Michele; Cavallini, Gabriella; Junquera, Concepción; Goya, Gerardo F; Raffa, Vittoria

    2017-04-01

    The only clinically approved alternative to autografts for treating large peripheral nerve injuries is the use of synthetic nerve guidance conduits (NGCs), which provide physical guidance to the regenerating stump and limit scar tissue infiltration at the injury site. Several lines of evidence suggest that a potential future strategy is to combine NGCs with cellular or molecular therapies to deliver growth factors that sustain the regeneration process. However, growth factors are expensive and have a very short half-life; thus, the combination approach has not been successful. In the present paper, we proposed the immobilization of growth factors (GFs) on magnetic nanoparticles (MNPs) for the time- and space-controlled release of GFs inside the NGC. We tested the particles in a rat model of a peripheral nerve lesion. Our results revealed that the injection of a cocktail of MNPs functionalized with nerve growth factor (NGF) and with vascular endothelial growth factor (VEGF) strongly accelerate the regeneration process and the recovery of motor function compared to that obtained using the free factors. Additionally, we found that injecting MNPs in the NGC is safe and does not impair the regeneration process, and the MNPs remain in the conduit for weeks. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Scaffolds from alternating block polyurethanes of poly(ɛ-caprolactone) and poly(ethylene glycol) with stimulation and guidance of nerve growth and better nerve repair than autograft.

    PubMed

    Niu, Yuqing; Li, Linjing; Chen, Kevin C; Chen, Feiran; Liu, Xiangyu; Ye, Jianfu; Li, Wei; Xu, Kaitian

    2015-07-01

    Nerve repair scaffolds from novel alternating block polyurethanes (PUCL-alt-PEG) based on PCL and PEG without additional growth factors or proteins were prepared by a particle leaching method. The scaffolds have pore size 10-20 µm and porosity 92%. Mechanical tests showed that the polyurethane scaffolds have maximum loads of 5.97 ± 0.35 N and maximal stresses of 8.84 ± 0.5 MPa. Histocompatiblity of the nerve repair scaffolds was tested in a SD rat model for peripheral nerve defect treatment. Two types of treatments including PUCL-alt-PEG scaffolds and autografts were compared in rat model. After 32 weeks, bridging of a 12 mm defect gap by the regenerated nerve was observed in all rats. The nerve regeneration was systematically characterized by sciatic function index (SFI), electrophysiology, histological assessment including HE staining, immunohistochemistry, ammonia sliver staining, Masson's trichrome staining and TEM observation. Results revealed that nerve repair scaffolds from PUCL-alt-PEG exhibit better regeneration effects compared to autografts. Electrophysiological recovery was seen in 90% and 87% of rats in PUCL-alt-PEG and autograft groups respectively. Biodegradation in vitro and in vivo shows good degradation match of PUCL-alt-PEG scaffolds with nerve regeneration. It demonstrates that plain nerve repair scaffolds from PUCL-alt-PEG biomaterials can achieve peripheral nerve regeneration satisfactorily.

  17. Plasticity in rat uterine sympathetic nerves: the role of TrkA and p75 nerve growth factor receptors

    PubMed Central

    Richeri, Analía; Bianchimano, Paola; Mármol, Nelson M; Viettro, Lorena; Cowen, Timothy; Brauer, M Mónica

    2005-01-01

    Uterine sympathetic innervation undergoes profound remodelling in response to physiological and experimental changes in the circulating levels of sex hormones. It is not known, however, whether this plasticity results from changes in the innervating neurons, the neuritogenic properties of the target tissue or both. Using densitometric immunohistochemistry, we analysed the effects of prepubertal chronic oestrogen treatment (three subcutaneous injections of 20 µg of β-oestradiol 17-cypionate on days 25, 27 and 29 after birth), natural peripubertal transition and late pregnancy (19–20 days post coitum) on the levels of TrkA and p75 nerve growth factor receptors in uterine-projecting sympathetic neurons of the thoraco-lumbar paravertebral sympathetic chain (T7–L2) identified using the retrograde tracer Fluorogold. For comparative purposes, levels of TrkA and p75 were assessed in the superior cervical ganglion (SCG) following prepubertal chronic oestrogen treatment. These studies showed that the vast majority of uterine-projecting neurons expressed both TrkA and p75. Both prepubertal chronic oestrogen treatment and the peripubertal transition increased the ratio p75 to TrkA in uterine-projecting neurons, whereas pregnancy elicited the opposite effect. Prepubertal chronic oestrogen treatment had no effects on levels of TrkA or p75 in sympathetic neurons of the SCG. Taken together, our data suggest that neurotrophin receptor-mediated events may contribute to regulate sex hormone-induced plasticity in uterine sympathetic nerves, and are in line with the idea that, in vivo, plasticity in uterine nerves involves changes in both the target and the innervating neurons. PMID:16050899

  18. Epidermal growth factor-induced cellular invasion requires sphingosine-1-phosphate/sphingosine-1-phosphate 2 receptor-mediated ezrin activation

    PubMed Central

    Orr Gandy, K. Alexa; Adada, Mohamad; Canals, Daniel; Carroll, Brittany; Roddy, Patrick; Hannun, Yusuf A.; Obeid, Lina M.

    2013-01-01

    Ezrin, radixin, and moesin (ERM) proteins link cortical actin to the plasma membrane and coordinate cellular events that require cytoskeletal rearrangement, including cell division, migration, and invasion. While ERM proteins are involved in many important cellular events, the mechanisms regulating their function are not completely understood. Our laboratory previously identified reciprocal roles for the sphingolipids ceramide and sphingosine-1-phosphate (S1P) in the regulation of ERM proteins. We recently showed that ceramide-induced activation of PP1α leads to dephosphorylation and inactivation of ERM proteins, while S1P results in phosphorylation and activation of ERM proteins. Following these findings, we aimed to examine known inducers of the SK/S1P pathway and evaluate their ability to regulate ERM proteins. We examined EGF, a known inducer of the SK/S1P pathway, for its ability to regulate the ERM family of proteins. We found that EGF induces ERM c-terminal threonine phosphorylation via activation of the SK/S1P pathway, as this was prevented by siRNA knockdown or pharmacological inhibition of SK. Using pharmacological, as well as genetic, knockdown approaches, we determined that EGF induces ERM phosphorylation via activation of S1PR2. In addition, EGF led to cell polarization in the form of lamellipodia, and this occurred through a mechanism involving S1PR2-mediated phosphorylation of ezrin T567. EGF-induced cellular invasion was also found to be dependent on S1PR2-induced T567 ezrin phosphorylation, such that S1PR2 antagonist, JTE-013, and expression of a dominant-negative ezrin mutant prevented cellular invasion toward EGF. In this work, a novel mechanism of EGF-stimulated invasion is unveiled, whereby S1P-mediated activation of S1PR2 and phosphorylation of ezrin T567 is required.—Orr Gandy, K. A., Adada, M., Canals, D., Carroll, B., Roddy, P., Hannun, Y. A., Obeid, L. M. Epidermal growth factor-induced cellular invasion requires sphingosine-1-phosphate

  19. Enhancement of sciatic nerve regeneration after vascular endothelial growth factor (VEGF) gene therapy.

    PubMed

    Pereira Lopes, F R; Lisboa, B C G; Frattini, F; Almeida, F M; Tomaz, M A; Matsumoto, P K; Langone, F; Lora, S; Melo, P A; Borojevic, R; Han, S W; Martinez, A M B

    2011-10-01

    Recent studies have emphasized the beneficial effects of the vascular endothelial growth factor (VEGF) on neurone survival and Schwann cell proliferation. VEGF is a potent angiogenic factor, and angiogenesis has long been recognized as an important and necessary step during tissue repair. Here, we investigated the effects of VEGF on sciatic nerve regeneration. Using light and electron microscopy, we evaluated sciatic nerve regeneration after transection and VEGF gene therapy. We examined the survival of the neurones in the dorsal root ganglia and in lumbar 4 segment of spinal cord. We also evaluated the functional recovery using the sciatic functional index and gastrocnemius muscle weight. In addition, we evaluated the VEGF expression by immunohistochemistry. Fluorescein isothiocyanate-dextran (FITC-dextran) fluorescence of nerves and muscles revealed intense staining in the VEGF-treated group. Quantitative analysis showed that the numbers of myelinated fibres and blood vessels were significantly higher in VEGF-treated animals. VEGF also increased the survival of neurone cell bodies in dorsal root ganglia and in spinal cord. The sciatic functional index and gastrocnemius muscle weight reached significantly higher values in VEGF-treated animals. We demonstrate a positive relationship between increased vascularization and enhanced nerve regeneration, indicating that VEGF administration can support and enhance the growth of regenerating nerve fibres, probably through a combination of angiogenic, neurotrophic and neuroprotective effects. © 2011 The Authors. Neuropathology and Applied Neurobiology © 2011 British Neuropathological Society.

  20. Nerve growth factor and epidermal growth factor stimulate clusterin gene expression in PC12 cells.

    PubMed Central

    Gutacker, C; Klock, G; Diel, P; Koch-Brandt, C

    1999-01-01

    Clusterin (apolipoprotein J) is an extracellular glycoprotein that might exert functions in development, cell death and lipid transport. Clusterin gene expression is elevated at sites of tissue remodelling, such as differentiation and apoptosis; however, the signals responsible for this regulation have not been identified. We use here the clusterin gene as a model system to examine expression in PC12 cells under the control of differentiation and proliferation signals produced by nerve growth factor (NGF) and by epidermal growth factor (EGF) respectively. NGF induced clusterin mRNA, which preceded neurite outgrowth typical of neuronal differentiation. EGF also activated the clusterin mRNA, demonstrating that both proliferation and differentiation signals regulate the gene. To localize NGF- and EGF-responsive elements we isolated the clusterin promoter and tested it in PC12 cell transfections. A 2.5 kb promoter fragment and two 1.5 and 0.3 kb deletion mutants were inducible by NGF and EGF. The contribution to this response of a conserved activator protein 1 (AP-1) motif located in the 0.3 kb fragment was analysed by mutagenesis. The mutant promoter was not inducible by NGF or EGF, which identifies the AP-1 motif as an element responding to both factors. Binding studies with PC12 nuclear extracts showed that AP-1 binds to this sequence in the clusterin promoter. These findings suggest that NGF and EGF, which give differential gene regulation in PC12 cells, resulting in neuronal differentiation and proliferation respectively, use the common Ras/extracellular signal-regulated kinase/AP-1 signalling pathway to activate clusterin expression. PMID:10215617

  1. Purinergic signalling underlies transforming growth factor‐β‐mediated bladder afferent nerve hyperexcitability

    PubMed Central

    Gonzalez, Eric J.; Heppner, Thomas J.; Nelson, Mark T.

    2016-01-01

    Key points The sensory components of the urinary bladder are responsible for the transduction of bladder filling and are often impaired with neurological injury or disease.Elevated extracellular ATP contributes, in part, to bladder afferent nerve hyperexcitability during urinary bladder inflammation or irritation.Transforming growth factor‐β1 (TGF‐β1) may stimulate ATP release from the urothelium through vesicular exocytosis mechanisms with minimal contribution from pannexin‐1 channels to increase bladder afferent nerve discharge.Bladder afferent nerve hyperexcitability and urothelial ATP release with CYP‐induced cystitis is decreased with TGF‐β inhibition.These results establish a causal link between an inflammatory mediator, TGF‐β, and intrinsic signalling mechanisms of the urothelium that may contribute to the altered sensory processing of bladder filling. Abstract The afferent limb of the micturition reflex is often compromised following bladder injury, disease and inflammatory conditions. We have previously demonstrated that transforming growth factor‐β (TGF‐β) signalling contributes to increased voiding frequency and decreased bladder capacity with cystitis. Despite the functional presence of TGF‐β in bladder inflammation, the precise mechanisms of TGF‐β mediating bladder dysfunction are not yet known. Thus, the present studies investigated the sensory components of the urinary bladder that may underlie the pathophysiology of aberrant TGF‐β activation. We utilized bladder–pelvic nerve preparations to characterize bladder afferent nerve discharge and the mechanisms of urothelial ATP release with distention. Our findings indicate that bladder afferent nerve discharge is sensitive to elevated extracellular ATP during pathological conditions of urinary bladder inflammation or irritation. We determined that TGF‐β1 may increase bladder afferent nerve excitability by stimulating ATP release from the urothelium via vesicular

  2. Complementary Effects of Two Growth Factors in Multifunctionalized Silk Nanofibers for Nerve Reconstruction

    PubMed Central

    Jose, Rodrigo R.; Vigneron, Pascale; Bresson, Damien; Fitzpatrick, Vincent; Marin, Frédéric; Kaplan, David L.; Egles, Christophe

    2014-01-01

    With the aim of forming bioactive guides for peripheral nerve regeneration, silk fibroin was electrospun to obtain aligned nanofibers. These fibers were functionalized by incorporating Nerve Growth Factor (NGF) and Ciliary NeuroTrophic Factor (CNTF) during electrospinning. PC12 cells grown on the fibers confirmed the bioavailability and bioactivity of the NGF, which was not significantly released from the fibers. Primary neurons from rat dorsal root ganglia (DRGs) were grown on the nanofibers and anchored to the fibers and grew in a directional fashion based on the fiber orientation, and as confirmed by growth cone morphology. These biofunctionalized nanofibers led to a 3-fold increase in neurite length at their contact, which was likely due to the NGF. Glial cell growth, alignment and migration were stimulated by the CNTF in the functionalized nanofibers. Organotypic culture of rat fetal DRGs confirmed the complementary effect of both growth factors in multifunctionalized nanofibers, which allowed glial cell migration, alignment and parallel axonal growth in structures resembling the ‘bands of Bungner’ found in situ. Graftable multi-channel conduits based on biofunctionalized aligned silk nanofibers were developed as an organized 3D scaffold. Our bioactive silk tubes thus represent new options for a biological and biocompatible nerve guidance conduit. PMID:25313579

  3. Rita Levi-Montalcini: the discovery of nerve growth factor and modern neurobiology.

    PubMed

    Aloe, Luigi

    2004-07-01

    The remarkable accomplishments in developmental neurobiology within the past 60 years have depended on two things: (i) a succession of original histochemical and immunohistochemical methodologies for identifying pathways in the nervous system with increasing precision and sensitivity, and (ii) the discovery of growth factors for neurons. Growth factors are naturally occurring, essential biological mediators that promote cell growth, differentiation, survival and function in specific nerve cell populations. The discovery of nerve growth factor (NGF) by Rita Levi-Montalcini in the 1950s represents an important milestone in the processes that led to modern cell biology. NGF was the first growth factor identified, for its action on the morphological differentiation of neural-crest-derived nerve cells. Later, its effect on neuronal cells of the peripheral and central nervous systems, and on several non-neuronal cells was also determined. Thus, Levi-Montalcini's work on NGF represents, as acknowledged by the Nobel Prize Assembly in its press release of 13 October 1986, "a fascinating example of how a skilled observer can create a concept out of apparent chaos".

  4. Nerve Growth Factor Effects on the Immune System

    DTIC Science & Technology

    1989-12-19

    growth factor protein, NGF, has been shown to play a physiologic role in the development and regeneration of the peripheral nervous system, acting on...determine the effects of NGF on lymphocytes with an aim to understanding the physiological and developmental role of NGF in the immune system. Our approaches...Thorpe and Perez-Polo, 1987). Although the maximal response of these cells occurs at levels significantly above those considered physiological it is

  5. Mesenchymal stem cells modified with nerve growth factor improve recovery of the inferior alveolar nerve after mandibular distraction osteogenesis in rabbits.

    PubMed

    Wang, L; Zhao, Y; Cao, J; Yang, X; Lei, D

    2015-03-01

    Distraction osteogenesis is widely used in the treatment of bony deformities and defects. However, injury to the inferior alveolar nerve is a concern. Our aim was to investigate the feasibility of using lentiviral-mediated human nerve growth factor beta (hNGFβ) of the inferior alveolar nerve in mandibular distraction osteogenesis in rabbits. To achieve this, mesenchymal stem cells (MSC) from the bone marrow of rabbit mandibles were isolated and genetically engineered using recombinant lentiviral vector containing hNGFβ. Twenty New Zealand white rabbits underwent mandibular distraction osteogenesis, and 5 million MSC transduced with hNGFβ-vector or control vector were transplanted around the nerve in the gap where the bone had been fractured during the operation (n=10 in each group). After gradual distraction, samples of the nerve were harvested for histological and histomorphometric analysis. We found that the genetically engineered MSC transduced by the lentiviral vector were able to secrete hNGFβ at physiologically relevant concentrations as measured by ELISA. Histological examination of the nerve showed more regenerating nerve fibres and less myelin debris in the group in which hNGFβ-modified MSC had been implanted than in the control group. Histomorphometric analysis of the nerve showed increased density of myelinated fibres in the group in which hNGFβ-modified MSC had been implanted than in the control group. The data suggest that implantation of hNGFβ-modified MSC can accelerate the morphological recovery of the inferior alveolar nerve during mandibular distraction osteogenesis in rabbits. The use of lentiviral-mediated gene treatment to deliver hNGFβ through MSC may be a promising way of minimising injury to the nerve. Copyright © 2015 The British Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.

  6. Nerve terminal growth remodels neuromuscular synapses in mice following regeneration of the postsynaptic muscle fiber.

    PubMed

    Li, Yue; Thompson, Wesley J

    2011-09-14

    Muscle fibers degenerate and regenerate in response to contractile damage, during aging, and in various muscle diseases that weaken the fibers. It is known that degeneration and regeneration of the segment of the postsynaptic fiber produces dramatic alterations in the neuromuscular junction (NMJ) that forms on the regenerated fiber, but the mechanisms here are incompletely understood. We have used a laser microbeam to damage the postsynaptic fibers at individual NMJs in the sternomastoid muscle of living young adult mice and then followed the synapses vitally over time using fluorescent proteins expressed in motor neurons and glial cells and staining of postsynaptic acetylcholine receptors. We find, in contrast to previous reports, that the mouse nerve terminal retains contact with the synaptic basal lamina marked by cholinesterase staining even in the absence of the target, showing that this terminal does not require a continuous supply of target-derived molecules for its maintenance. Thus, remodeling of the nerve terminal during the period of target absence does not explain the subsequent changes in the new NMJ. Rather, we see that the synapse becomes altered as the new fiber segment regenerates. Mechanisms for remodeling the synapse include failure of the regenerating muscle fiber to contact the old basal lamina and nerve terminal, growth of the nerve terminal and its glia toward the regenerating fiber, and remodeling of the initial contact as the nerve terminal becomes varicose.

  7. Combined treatment with FK506 and nerve growth factor for spinal cord injury in rats.

    PubMed

    Chen, Guang; Zhang, Zhen; Wang, Shouyu; Lv, Decheng

    2013-10-01

    Following spinal cord injury in rats, FK506 is able to protect local nerve tissue, promote neural regeneration, reduce neuronal apoptosis and accelerate the recovery of spinal cord functions. Nerve growth factor (NGF) is important in the regulation of central and peripheral nerve cell regeneration, growth differentiation and functions. Previous studies have shown that FK506 and NGF exhibit a synergistic effect in the treatment of peripheral nerve injury; however, it remains unclear whether the synergistic effect is present in the treatment of spinal cord injury. In this study, we combined FK506 and NGF for the treatment of spinal cord injury in rats. The NF200 protein expression in rats with spinal cord injury was determined using immunohistochemical staining and NF200 mRNA expression levels were observed using the reverse transcription-polymerase chain reaction method. The restoration of spinal cord functions was evaluated using the Basso, Beattie and Bresnahan score. The results demonstrated that the combined treatment significantly enhanced the expression of NF200 and improved spinal cord functions compared with the results of the single treatment. Our experimental observations indicated that FK506 and NGF exhibit a synergistic effect in the treatment of spinal cord injury in rats and that the combined treatment may effectively promote neural regeneration and functional recovery in rats following spinal cord injury.

  8. Molecular cloning of a human gene that is a member of the nerve growth factor family

    SciTech Connect

    Jones, K.R.; Reichardt, L.F. )

    1990-10-01

    Cell death within the developing vertebrate nervous system is regulated in part by interactions between neurons and their innervation targets that are mediated by neurotrophic factors. These factors also appear to have a role in the maintenance of the adult nervous system. Two neurotrophic factors, nerve growth factor and brain-derived neurotrophic factor, share substantial amino acid sequence identity. The authors have used a screen that combines polymerase chain reaction amplification of genomic DNA and low-stringency hybridization with degenerate oligonucleotides to isolate human BDNF and a human gene, neurotrophin-3, that is closely related to both nerve growth factor and brain-derived neurotrophic factor. mRNA products of the brain-derived neurotrophic factor and neurotrophin-3 genes were detected in the adult human brain, suggesting that these proteins are involved in the maintenance of the adult nervous system. Neurotrophin-3 is also expected to function in embryonic neural development.

  9. Nerve growth factor corrects developmental impairments of basal forebrain cholinergic neurons in the trisomy 16 mouse.

    PubMed Central

    Corsi, P; Coyle, J T

    1991-01-01

    The trisomy 16 (Ts16) mouse, which shares genetic and phenotypic homologies with Down syndrome, exhibits impaired development of the basal forebrain cholinergic system. Basal forebrains obtained from Ts16 and euploid littermate fetuses at 15 days of gestation were dissociated and cultured in completely defined medium, with cholinergic neurons identified by choline acetyltransferase (ChAT) immunoreactivity. The Ts16 cultures exhibited fewer ChAT-immunoreactive neurons, which were smaller and emitted shorter, smoother, and more simplified neurites than those from euploid littermates. Whereas the addition of beta-nerve growth factor (100 ng/ml) augmented the specific activity of ChAT and neuritic extension for both Ts16 and euploid cholinergic neurons, only Ts16 cultures exhibited an increase in the number and size of ChAT-immunoreactive neurons. Furthermore, Ts16 ChAT-immunoreactive neurites formed varicosities only in the presence of beta-nerve growth factor. Images PMID:2000385

  10. Soft Graphene Nanofibers Designed for the Acceleration of Nerve Growth and Development.

    PubMed

    Feng, Zhang-Qi; Wang, Ting; Zhao, Bin; Li, Jiacheng; Jin, Lin

    2015-11-04

    Soft graphene nanofibers with recoverable electrical conductivity and excellent physicochemical stability are prepared by a controlled assembly technique. By using the soft graphene nanofibers for cellular electrical stimulation, the common inhibitory effect of long-term electrical stimulation on nerve growth and development is avoided, which usually happens with traditional 2D conductive materials. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Changes in myelin sheath thickness and internode geometry in the rabbit phrenic nerve during growth.

    PubMed Central

    Friede, R L; Brzoska, J; Hartmann, U

    1985-01-01

    The rabbit phrenic nerve was studied at seven phases of growth from the newborn to the adult to determine the length of the nerve fibres, the length of the internodes, the fibre calibre, the geometric proportions of the internodes and the thickness of the myelin sheaths. The elongation of the internodes corresponded precisely to the elongation of the nerve, indicating a constant number of approximately 140 internodes per fibre, each internode elongating commensurate with body growth. Internode elongation was accompanied by increases in fibre calibre, but these parameters did not change in precise proportion. The internodes of thick fibres were relatively short for calibre, as defined by the length/diameter quotient. This trend of foreshortening changed during growth. Sheath thickness, defined by the quotient axon diameter/fibre diameter, was determined with a computer-assisted method. Fibres of young rabbits had relatively thin sheaths for axon calibre, compared with adult rabbits. The changes in sheath thickness corresponded to the changes in internode geometry. This was consistent with previous studies showing that elongation or foreshortening of an internode of a given calibre has a slight, but definite effect on the thickness of its myelin sheath. PMID:3870716

  12. Human Adenomyosis Endometrium Stromal Cells Secreting More Nerve Growth Factor: Impact and Effect.

    PubMed

    Li, Yan; Zou, Shien; Xia, Xian; Zhang, Shaofen

    2015-09-01

    Abnormal expression of nerve growth factor (NGF) was found in adenomyosis (AM). We collected AM foci from patients and eutopic endometrium from non-AM controls. Endometrium stromal cells (ESCs) were cultured. Different levels of 17β-estradiol, tumor necrosis factor (TNF), CoCl2, and H2O2 were added to the culture system separately, then the expression level of NGF in ESCs was detected. After adding different levels of NGF, the proliferation and apoptosis of ESCs and aromatase expression were detected. We found that 17β-estradiol promoted NGF production in AM ESCs but not in control ESCs; TNF promoted NGF production in both AM and control ESCs; and CoCl2 inhibited NGF production in control ESCs, but had no effect in AM ESCs. Nerve growth factor promoted the proliferation and synthesis of aromatase in AM ESCs. In conclusion, locally increased estrogen levels and inflammation may cause increased NGF production in the uterus of patients with AM. Nerve growth factor stimulated the proliferation and increased aromatase expression of ESCs from AM foci, suggesting NGF might contribute to the pathology and etiology of AM.

  13. Immunity to nerve growth factor and the effect on motor unit reinnervation in the rabbit.

    PubMed

    Finkelstein, D I; Luff, A R; Schuijers, J A

    1992-05-01

    The trophic effects of nerve growth factor (NGF) on sympathetic, peripheral afferent, and other neural crest-derived cells have been intensively investigated. More recently, NGF has been shown to have an influence on motoneurons. This study was undertaken to investigate whether NGF had any influence on the mechanical or histological properties of reinnervated motor units. Three groups of rabbits were used: normal rabbits, rabbits in which the nerve to medial gastrocnemius (MG) was cut and allowed to reinnervate for 56 days, and rabbits in which the MG nerve reinnervated in the presence of immunity to NGF. Immunity to NGF did not affect the ability of motor axons to reinnervate a muscle, nor were the contractile characteristics of the motor units altered. The size of horseradish peroxidase-labeled motoneurons was not influenced by immunization against NGF; however, the distribution of afferent neuron sizes was altered. Conduction velocity of motor axons proximal to the neuroma was significantly faster after immunization against NGF. Transection and subsequent reinnervation by a peripheral nerve normally causes an increase in myelin thickness proximal to the neuroma. However, immunization against NGF appeared to decrease the magnitude of myelin thickening. It was concluded that immunization against NGF affects motor axonal conduction velocity via an influence on the neural crest-derived Schwann cells.

  14. Chondroitinase C Selectively Degrades Chondroitin Sulfate Glycosaminoglycans that Inhibit Axonal Growth within the Endoneurium of Peripheral Nerve

    PubMed Central

    2016-01-01

    The success of peripheral nerve regeneration is highly dependent on the regrowth of axons within the endoneurial basal lamina tubes that promote target-oriented pathfinding and appropriate reinnervation. Restoration of nerve continuity at this structural level after nerve transection injury by direct repair and nerve grafting remains a major surgical challenge. Recently, biological approaches that alter the balance of growth inhibitors and promoters in nerve have shown promise to improve appropriate axonal regeneration and recovery of peripheral nerve function. Chondroitin sulfate proteoglycans (CSPGs) are known inhibitors of axonal growth. This growth inhibition is mainly associated with a CSPG's glycosaminoglycan chains. Enzymatic degradation of these chains with chondroitinase eliminates this inhibitory activity and, when applied in vivo, can improve the outcome of nerve repair. To date, these encouraging findings were obtained with chondroitinase ABC (a pan-specific chondroitinase). The aim of this study was to examine the distribution of CSPG subtypes in rodent, rabbit, and human peripheral nerve and to test more selective biological enzymatic approaches to improve appropriate axonal growth within the endoneurium and minimize aberrant growth. Here we provide evidence that the endoneurium, but not the surrounding epineurium, is rich in CSPGs that have glycosaminoglycan chains readily degraded by chondroitinase C. Biochemical studies indicate that chondroitinase C has degradation specificity for 6-sulfated glycosaminoglycans found in peripheral nerve. We found that chondroitinase C degrades and inactivates inhibitory CSPGs within the endoneurium but not so much in the surrounding nerve compartments. Cryoculture bioassays (neurons grown on tissue sections) show that chondroitinase C selectively and significantly enhanced neuritic growth associated with the endoneurial basal laminae without changing growth-inhibiting properties of the surrounding epineurium

  15. Nerve growth factor (NGF) and diabetic neuropathy in the rat: morphological investigations of the sural nerve, dorsal root ganglion, and spinal cord.

    PubMed

    Unger, J W; Klitzsch, T; Pera, S; Reiter, R

    1998-09-01

    A number of functions for nerve growth factor (NGF) have been described over the past years, including its role for neuronal function and regeneration during toxic or metabolic neuropathies. In order to further assess the effects of NGF on the somatosensory system in diabetic neuropathy, the sural nerve, dorsal root ganglia (DRG), and dorsal horn of the spinal cord were investigated by morphological and quantitative methods in rats after 12 weeks of uncontrolled streptozotocin-induced diabetes mellitus. The results from our study suggest a twofold effect of NGF: (1) In sural nerve treatment with NGF (0.1 or 0.5 mg/kg) for 12 weeks was able to reverse distinct diabetes-related alterations in myelinated nerve fiber morphology, such as myelin thickness. These changes occurred in the entire myelinated population of sensory nerves and were not restricted to nociceptive nerve fibers. (2) The NGF effect on neurotransmitters of the sensory, nociceptive system was reflected by increased CGRP and substance P content in the DRG and in the dorsal horn of the spinal cord. No change of trkA receptor immunostaining was seen in DRGs of diabetic rats; however, a reduction of trkA immunoreactivity of DRG neurons was noted after long-term NGF treatment of healthy controls. The data demonstrate that NGF regulates a number of neuronal parameters along peripheral and central parts of the somatosensory pathway in the adult. This neurotrophic support may be essential for inducing functionally significant regenerative mechanisms in diabetic neuropathy.

  16. Effect of local administration of platelet-derived growth factor B on functional recovery of peripheral nerve regeneration: A sciatic nerve transection model

    PubMed Central

    Golzadeh, Atefeh; Mohammadi, Rahim

    2016-01-01

    Background: Effects of platelet-derived growth factor B (PDGF-B) on peripheral nerve regeneration was studied using a rat sciatic nerve transection model. Materials and Methods: Forty-five male, white Wistar rats were divided into three experimental groups (n = 15), randomly: Normal control group (NC), silicon group (SIL), and PDGF-B treated group (SIL/PDGF). In NC group, left sciatic nerve was exposed through a gluteal muscle incision and after homeostasis muscle was sutured. In the SIL group, the left sciatic nerve was exposed in the same way and transected proximal to tibio-peroneal bifurcation leaving a 10-mm gap. Proximal and distal stumps were each inserted into a silicone conduit and filled with 10 μL phosphate buffered solution. In SIL/PDGF group, the silicon conduit was filled with 10 μL PDGF-B (0.5 ng/mL). Each group was subdivided into three subgroups of five and were studied in 4, 8, 12 weeks after surgery. Results: Behavioral testing, sciatic nerve functional study, gastrocnemius muscle mass, and histomorphometric studies showed earlier regeneration of axons in SIL/PDGF than in SIL group (P < 0.05). Conclusion: Local administration of PDGF-B combined with silicon grafting could accelerate functional recovery and may have clinical implications for the surgical management of patients after facial nerve transection. PMID:27274342

  17. Effects of locally applied nerve growth factor to the inferior alveolar nerve histology in a rabbit model of mandibular distraction osteogenesis.

    PubMed

    Wang, L; Zhao, Y; Cheng, X; Yang, Y; Liu, G; Ma, Q; Shang, H; Tian, L; Lei, D

    2009-01-01

    Distraction osteogenesis (DO) is widely used in deformities and defects of the craniofacial bone. Accelerating inferior alveolar nerve (IAN) recovery would aid the process. Nerve growth factor (NGF) plays a vital role in peripheral nerve regeneration. In this study, the ability of locally applied human NGF beta (hNGFbeta) to enhance the morphological recovery of the IAN in a rabbit model of mandibular DO was studied. Rabbits underwent bilateral DO with a rate of 0.5mm per 12h. Two doses of 40 microg hNGFbeta in buffer were injected into callus at the beginning the of consolidation time. The contralateral side received injections of placebo. Rabbits were killed at 14 and 28 days. IAN specimens were subjected to histological and histomorphometric analysis. In both 14 and 28 days consolidation experiments, nerve histological analysis showed less degeneration and more regeneration in nerve fibers on the hNGFbeta treated side than the control side. Histomorphometric analysis showed that the myelinated fiber density on the hNGFbeta treated side was significantly higher than on the control side (p<0.01). The data indicate that locally applied hNGFbeta can accelerate the morphological recovery of the IAN and may play a role in reducing nerve injury in mandibular DO clinically.

  18. The effects of gradients of nerve growth factor immobilized PCLA scaffolds on neurite outgrowth in vitro and peripheral nerve regeneration in rats.

    PubMed

    Tang, Shuo; Zhu, Jixiang; Xu, Yangbin; Xiang, Andy Peng; Jiang, Mei Hua; Quan, Daping

    2013-09-01

    Introducing concentration gradients of nerve growth factor (NGF) into conduits for repairing of peripheral nerve injury is crucial for nerve regeneration and guidance. Herein, combining differential adsorption of NGF/silk fibroin (SF) coating, the gradient of NGF-immobilized membranes (G-Ms) and nanofibrous nerve conduits (G-nNCs) were successfully fabricated. The efficacy of NGF gradients was confirmed by a quantitative comparison of dorsal root ganglia (DRG) neurite outgrowth on the G-Ms or uniform NGF-immobilized membranes (U-Ms). Significantly, the neurite turning ratio was 0.48 ± 0.11 for G-M group, but it was close to zero for U-M group. The neurite length of DRGs in the middle of the G-Ms was significantly longer than that of U-M group, even though the average NGF concentration was approximated. Furthermore, 12 weeks after implantation in rats with a 14 mm gap of sciatic nerve injury, G-nNCs achieved satisfying outcomes of nerve regeneration associated with morphological and functional improvements, which was superior to that of the uniform NGF-immobilized nNCs (U-nNCs). Sciatic function index (SFI), compound muscle action potentials (CMAPs), total number of myelinated nerve fibers, thickness of myelin sheath were similar for the G-nNCs and autografts, with the G-nNCs having a higher density of axons than the autografts. Our results demonstrated the significant role of introducing NGF gradients into scaffolds in promoting nerve regeneration. Copyright © 2013 Elsevier Ltd. All rights reserved.

  19. Angelica injection promotes peripheral nerve structure and function recovery with increased expressions of nerve growth factor and brain derived neurotrophic factor in diabetic rats.

    PubMed

    Li, Ruilin; Zhang, Junjian; Zhang, Lei; Cui, Qin; Liu, Hui

    2010-08-01

    Several nervous system injury models, such as sciatic crush and chronic cerebral hypoperfusion have been well studied in terms of neuroprotective effect of angelica injection. However, definitive experimental studies are lacking on diabetic peripheral neuropathy (DPN). This study sought to investigate the effects of angelica injection on DPN in type 1 diabetic rats. Diabetes was induced by single intraperitoneal injection of streptozotocin (STZ). To examine whether DPN model succeeded, tail-flick latency (TFL) and motor nerve conduction velocity (MNCV) were measured at 6 weeks after diabetes induction. Then, diabetic rats were treated with high- and low-dose angelica injection for 4 weeks. TFL, MNCV, morphology of sciatic nerve, myelinated nerve fiber density and the expressions of nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF) in soleus and sciatic nerve were measured at 10 weeks after diabetes induction. The results showed the TFL was significantly shortened (p<0.001) and the MNCV was reduced (p<0.01) in diabetic rats compared with normal control rats at 6 weeks after diabetes induction. The TFL was obviously prolonged and the MNCV was further reduced in diabetic control group at 10 weeks after diabetes induction. TFL, MNCV and morphology of sciatic nerve were remarkably ameliorated and myelinated nerve fiber density and the expressions of NGF and BDNF in soleus and sciatic nerve were increased in the angelica treatment groups. This study suggests angelica injection has potential therapeutic effects on DPN, and the mechanism might be related to direct increase in NGF expression and direct or indirect increase in BDNF expression.

  20. Secretion of Growth Hormone in Response to Muscle Sensory Nerve Stimulation

    NASA Technical Reports Server (NTRS)

    Grindeland, Richard E.; Roy, R. R.; Edgerton, V. R.; Gosselink, K. L.; Grossman, E. J.; Sawchenko, P. E.; Wade, Charles E. (Technical Monitor)

    1994-01-01

    Growth hormone (GH) secretion is stimulated by aerobic and resistive exercise and inhibited by exposure to actual or simulated (bedrest, hindlimb suspension) microgravity. Moreover, hypothalamic growth hormone-releasing factor (GRF) and preproGRF mRNA are markedly decreased in spaceflight rats. These observations suggest that reduced sensory input from inactive muscles may contribute to the reduced secretion of GH seen in "0 G". Thus, the aim of this study was to determine the effect of muscle sensory nerve stimulation on secretion of GH. Fed male Wistar rats (304 +/- 23 g) were anesthetized (pentobarbital) and the right peroneal (Pe), tibial (T), and sural (S) nerves were cut. Electrical stimulation of the distal (D) or proximal (P) ends of the nerves was implemented for 15 min. to mimic the EMG activity patterns of ankle extensor muscles of a rat walking 1.5 mph. The rats were bled by cardiac puncture and their anterior pituitaries collected. Pituitary and plasma bioactive (BGH) and immunoactive (IGH) GH were measured by bioassay and RIA.

  1. Nerve Growth Factor Promoter Activity Revealed in Mice Expressing Enhanced Green Fluorescent Protein

    PubMed Central

    Kawaja, Michael D.; Smithson, Laura J.; Elliott, Janet; Trinh, Gina; Crotty, Anne-Marie; Michalski, Bernadeta; Fahnestock, Margaret

    2012-01-01

    Nerve growth factor (NGF) and its precursor proNGF are perhaps the best described growth factors of the mammalian nervous system. There remains, however, a paucity of information regarding the precise cellular sites of proNGF/NGF synthesis. Here we report the generation of transgenic mice in which the NGF promoter controls the ectopic synthesis of enhanced green fluorescent protein (EGFP). These transgenic mice provide an unprecedented resolution of both neural cells (e.g., neocortical and hippocampal neurons) and non-neural cells (e.g., renal interstitial cells and thymic reticular cells) that display NGF promoter activity from postnatal development to adulthood. Moreover, the transgene is inducible by injury. At 2 days after sciatic nerve ligation, a robust population of EGFP-positive cells is seen in the proximal nerve stump. These transgenic mice offer novel insights into the cellular sites of NGF promoter activity and can be used as models for investigating the regulation of proNGF/NGF expression after injury. PMID:21456011

  2. Secretion of Growth Hormone in Response to Muscle Sensory Nerve Stimulation

    NASA Technical Reports Server (NTRS)

    Grindeland, Richard E.; Roy, R. R.; Edgerton, V. R.; Gosselink, K. L.; Grossman, E. J.; Sawchenko, P. E.; Wade, Charles E. (Technical Monitor)

    1994-01-01

    Growth hormone (GH) secretion is stimulated by aerobic and resistive exercise and inhibited by exposure to actual or simulated (bedrest, hindlimb suspension) microgravity. Moreover, hypothalamic growth hormone-releasing factor (GRF) and preproGRF mRNA are markedly decreased in spaceflight rats. These observations suggest that reduced sensory input from inactive muscles may contribute to the reduced secretion of GH seen in "0 G". Thus, the aim of this study was to determine the effect of muscle sensory nerve stimulation on secretion of GH. Fed male Wistar rats (304 +/- 23 g) were anesthetized (pentobarbital) and the right peroneal (Pe), tibial (T), and sural (S) nerves were cut. Electrical stimulation of the distal (D) or proximal (P) ends of the nerves was implemented for 15 min. to mimic the EMG activity patterns of ankle extensor muscles of a rat walking 1.5 mph. The rats were bled by cardiac puncture and their anterior pituitaries collected. Pituitary and plasma bioactive (BGH) and immunoactive (IGH) GH were measured by bioassay and RIA.

  3. Primary neuron culture for nerve growth and axon guidance studies in zebrafish (Danio rerio).

    PubMed

    Chen, Zheyan; Lee, Han; Henle, Steven J; Cheever, Thomas R; Ekker, Stephen C; Henley, John R

    2013-01-01

    Zebrafish (Danio rerio) is a widely used model organism in genetics and developmental biology research. Genetic screens have proven useful for studying embryonic development of the nervous system in vivo, but in vitro studies utilizing zebrafish have been limited. Here, we introduce a robust zebrafish primary neuron culture system for functional nerve growth and guidance assays. Distinct classes of central nervous system neurons from the spinal cord, hindbrain, forebrain, and retina from wild type zebrafish, and fluorescent motor neurons from transgenic reporter zebrafish lines, were dissociated and plated onto various biological and synthetic substrates to optimize conditions for axon outgrowth. Time-lapse microscopy revealed dynamically moving growth cones at the tips of extending axons. The mean rate of axon extension in vitro was 21.4±1.2 µm hr(-1) s.e.m. for spinal cord neurons, which corresponds to the typical ∼0.5 mm day(-1) growth rate of nerves in vivo. Fluorescence labeling and confocal microscopy demonstrated that bundled microtubules project along axons to the growth cone central domain, with filamentous actin enriched in the growth cone peripheral domain. Importantly, the growth cone surface membrane expresses receptors for chemotropic factors, as detected by immunofluorescence microscopy. Live-cell functional assays of axon extension and directional guidance demonstrated mammalian brain-derived neurotrophic factor (BDNF)-dependent stimulation of outgrowth and growth cone chemoattraction, whereas mammalian myelin-associated glycoprotein inhibited outgrowth. High-resolution live-cell Ca(2+)-imaging revealed local elevation of cytoplasmic Ca(2+) concentration in the growth cone induced by BDNF application. Moreover, BDNF-induced axon outgrowth, but not basal outgrowth, was blocked by treatments to suppress cytoplasmic Ca(2+) signals. Thus, this primary neuron culture model system may be useful for studies of neuronal development, chemotropic axon

  4. Nestin-Expressing Stem Cells Promote Nerve Growth in Long-Term 3-Dimensional Gelfoam®-Supported Histoculture

    PubMed Central

    Mii, Sumiyuki; Uehara, Fuminari; Yano, Shuya; Tran, Benjamin; Miwa, Shinji; Hiroshima, Yukihiro; Amoh, Yasuyuki; Katsuoka, Kensei; Hoffman, Robert M.

    2013-01-01

    We have previously reported that hair follicles contain multipotent stem cells which express nestin. The nestin-expressing cells form the hair follicle sensory nerve. In vitro, the nestin-expressing hair follicle cells can differentiate into neurons, Schwann cells, and other cell types. In the present study, the sciatic nerve was excised from transgenic mice in which the nestin promoter drives green fluorescent protein (ND-GFP mice). The ND-GFP cells of the sciatic nerve were also found to be multipotent as the ND-GFP cells in the hair follicle. When the ND-GFP cells in the mouse sciatic nerve cultured on Gelfoam® and were imaged by confocal microscopy, they were observed forming fibers extending the nerve. The fibers consisted of ND-GFP-expressing spindle cells, which co-expressed the neuron marker β-III tubulin, the immature Schwann-cell marker p75NTR and TrkB which is associated with neurons. The fibers also contain nestin-negative spherical cells expressing GFAP, a Schwann-cell marker. The β-III tubulin-positive fibers had growth cones on their tips expressing F-actin, indicating they are growing axons. When the sciatic nerve from mice ubiquitously expressing red fluorescent protein (RFP) was co-cultured on Gelfoam® with the sciatic nerve from ND-GFP transgenic mice, the interaction of nerves was observed. Proliferating nestin-expressing cells in the injured sciatic nerve were also observed in vivo. Nestin-expressing cells were also observed in posterior nerves but not in the spinal cord itself, when placed in 3-D Gelfoam® culture. The results of the present report suggest a critical function of nestin-expressing cells in peripheral nerve growth and regeneration. PMID:23840607

  5. Incorporation of Chitosan Microspheres into Collagen-Chitosan Scaffolds for the Controlled Release of Nerve Growth Factor

    PubMed Central

    Xiao, Wei; Qi, Fengyu; Huang, Jinghui; Luo, Zhuojing

    2014-01-01

    Background Artifical nerve scaffold can be used as a promising alternative to autologous nerve grafts to enhance the repair of peripheral nerve defects. However, current nerve scaffolds lack efficient microstructure and neurotrophic support. Methods Microsphere–Scaffold composite was developed by incorporating chitosan microspheres loaded with nerve growth factor (NGF–CMSs) into collagen-chitosan scaffolds (CCH) with longitudinally oriented microchannels (NGF–CMSs/CCH). The morphological characterizations, in vitro release kinetics study, neurite outgrowth assay, and bioactivity assay were evaluated. After that, a 15-mm-long sciatic nerve gap in rats was bridged by the NGF–CMSs/CCH, CCH physically absorbed NGF (NGF/CCH), CCH or nerve autograft. 16 weeks after implantation, electrophysiology, fluoro-gold retrograde tracing, and nerve morphometry were performed. Results The NGF–CMSs were evenly distributed throughout the longitudinally oriented microchannels of the scaffold. The NGF–CMSs/CCH was capable of sustained release of bioactive NGF within 28 days as compared with others in vitro. In vivo animal study demonstrated that the outcomes of NGF–CMSs/CCH were better than those of NGF/CCH or CCH. Conclusion Our findings suggest that incorporation of NGF–CMSs into the CCH may be a promising tool in the repair of peripheral nerve defects. PMID:24983464

  6. Expression of nerve growth factor receptor in paraffin-embedded soft tissue tumors.

    PubMed Central

    Perosio, P. M.; Brooks, J. J.

    1988-01-01

    Identification of growth factors and receptors in mesenchymal tumors may be crucial to understanding of growth regulation in sarcomas. During an immunohistochemical study of the expression of growth factors and receptors in human soft tissue tumors (STT), only 1 antisera capable of working in paraffin-embedded tissue was noted. A detailed study of 141 STT was undertaken to determine the frequency of expression of nerve growth factor receptor (NGF-R), its specificity and sensitivity for neural tumors, and the effect of fixation on detection. In normal mesenchymal tissue, only nerve sheath and perivascular staining was seen. No immunoreactivity was seen in many tumors including rhabdomyosarcoma, angiosarcoma, liposarcoma, Ewing's sarcoma, and alveolar soft part sarcoma. Less than 15% of tumors of smooth muscle, fibrous, or fibrohistiocytic origin showed immunoreactivity, usually focal. In contrast, a high frequency of immunoreactivity was noted in tumors of neural origin (74%). This included granular cell tumors (100%), Schwannoma/neurofibroma (91%), malignant Schwannoma (78%), neuroblastoma/neuroepithelioma (60%), and paraganglioma (57%). A high rate of reactivity was also seen in synovial sarcomas (80%), undifferentiated sarcomas (60%), and hemangiopericytomas (43%), suggesting a potential relationship to the neural phenotype. Among the neural tumors, Bouin's fixation was superior to formalin, suggesting that immunoreactivity for NGF-R is affected by fixation. This antibody may be a useful adjunct marker diagnostically. Images Figure 1 Figure 2 Figure 4 Figure 5 Figure 6 Figure 7 Figure 9 Figure 10 PMID:2456020

  7. Corneal sulfated glycosaminoglycans and their effects on trigeminal nerve growth cone behavior in vitro: roles for ECM in cornea innervation.

    PubMed

    Schwend, Tyler; Deaton, Ryan J; Zhang, Yuntao; Caterson, Bruce; Conrad, Gary W

    2012-12-13

    Sensory trigeminal nerve growth cones innervate the cornea in a highly coordinated fashion. The purpose of this study was to determine if extracellular matrix glycosaminoglycans (ECM-GAGs), including keratan sulfate (KS), dermatan sulfate (DS), and chondroitin sulfate A (CSA) and C (CSC), polymerized in developing eyefronts, may provide guidance cues to nerves during cornea innervation. Immunostaining using antineuron-specific-β-tubulin and monoclonal antibodies for KS, DS, and CSA/C was performed on eyefronts from embryonic day (E) 9 to E14 and staining visualized by confocal microscopy. Effects of purified GAGs on trigeminal nerve growth cone behavior were tested using in vitro neuronal explant cultures. At E9 to E10, nerves exiting the pericorneal nerve ring grew as tight fascicles, advancing straight toward the corneal stroma. In contrast, upon entering the stroma, nerves bifurcated repeatedly as they extended anteriorly toward the epithelium. KS was localized in the path of trigeminal nerves, whereas DS and CSA/C-rich areas were avoided by growth cones. When E10 trigeminal neurons were cultured on different substrates comprised of purified GAG molecules, their neurite growth cone behavior varied depending on GAG type, concentration, and mode of presentation (immobilized versus soluble). High concentrations of immobilized KS, DS, and CSA/C inhibited neurite growth to varying degrees. Neurites traversing lower, permissive concentrations of immobilized DS and CSA/C displayed increased fasciculation and decreased branching, whereas KS caused decreased fasciculation and increased branching. Enzymatic digestion of sulfated GAGs canceled their effects on trigeminal neurons. Data herein suggest that GAGs may direct the movement of trigeminal nerve growth cones innervating the cornea.

  8. Corneal Sulfated Glycosaminoglycans and Their Effects on Trigeminal Nerve Growth Cone Behavior In Vitro: Roles for ECM in Cornea Innervation

    PubMed Central

    Schwend, Tyler; Deaton, Ryan J.; Zhang, Yuntao; Caterson, Bruce; Conrad, Gary W.

    2012-01-01

    Purpose. Sensory trigeminal nerve growth cones innervate the cornea in a highly coordinated fashion. The purpose of this study was to determine if extracellular matrix glycosaminoglycans (ECM–GAGs), including keratan sulfate (KS), dermatan sulfate (DS), and chondroitin sulfate A (CSA) and C (CSC), polymerized in developing eyefronts, may provide guidance cues to nerves during cornea innervation. Methods. Immunostaining using antineuron-specific-β-tubulin and monoclonal antibodies for KS, DS, and CSA/C was performed on eyefronts from embryonic day (E) 9 to E14 and staining visualized by confocal microscopy. Effects of purified GAGs on trigeminal nerve growth cone behavior were tested using in vitro neuronal explant cultures. Results. At E9 to E10, nerves exiting the pericorneal nerve ring grew as tight fascicles, advancing straight toward the corneal stroma. In contrast, upon entering the stroma, nerves bifurcated repeatedly as they extended anteriorly toward the epithelium. KS was localized in the path of trigeminal nerves, whereas DS and CSA/C–rich areas were avoided by growth cones. When E10 trigeminal neurons were cultured on different substrates comprised of purified GAG molecules, their neurite growth cone behavior varied depending on GAG type, concentration, and mode of presentation (immobilized versus soluble). High concentrations of immobilized KS, DS, and CSA/C inhibited neurite growth to varying degrees. Neurites traversing lower, permissive concentrations of immobilized DS and CSA/C displayed increased fasciculation and decreased branching, whereas KS caused decreased fasciculation and increased branching. Enzymatic digestion of sulfated GAGs canceled their effects on trigeminal neurons. Conclusions. Data herein suggest that GAGs may direct the movement of trigeminal nerve growth cones innervating the cornea. PMID:23132805

  9. Vascular endothelial growth factor promotes anatomical and functional recovery of injured peripheral nerves in the avascular cornea

    PubMed Central

    Pan, Zan; Fukuoka, Shima; Karagianni, Natalia; Guaiquil, Victor H.; Rosenblatt, Mark I.

    2013-01-01

    Peripheral nerve injury is a major neurological disorder that can cause severe motor and sensory dysfunction. Neurogenic effects of vascular endothelial growth factor (VEGF) have been found in the central nervous system, and we examined whether VEGF could promote anatomical and functional recovery of peripheral nerves after injury using an avascular corneal nerve injury model. We found that VEGF enhanced neurite elongation in isolated trigeminal ganglion neurons in a dose-dependent manner. This effect was suppressed by neutralizing antibodies for VEGF receptor (VEGFR) 1 and 2 or neuropilin receptor 1 or by VEGFR2 inhibitors (SU 1498 and Ki 8751). In vivo, mice receiving sustained VEGF via implanted pellets showed increased corneal nerve regeneration after superficial injury compared with those receiving vehicle. VEGF injected subconjunctivally at the time of injury accelerated reinnervation, the recovery of mechanosensation, and epithelial wound healing. Endogenous VEGF expression was up-regulated in the corneal epithelium and stroma after wounding. Thus, VEGF can mediate peripheral neuron growth but requires the activation of multiple VEGF receptor types. In addition, VEGF can accelerate the return of sensory and trophic functions of damaged peripheral nerves. Wounding induces the expression of VEFG, which may modulate physiological nerve repair.—Pan, Z., Fukuoka, S., Karagianni, N., Guaiquil, V. H., Rosenblatt, M. I. Vascular endothelial growth factor promotes anatomical and functional recovery of injured peripheral nerves in the avascular cornea. PMID:23568776

  10. Increased severity of inflammation correlates with elevated expression of TRPV1 nerve fibers and nerve growth factor on interstitial cystitis/bladder pain syndrome.

    PubMed

    Liu, Bo-long; Yang, Fei; Zhan, Hai-lun; Feng, Zhi-ying; Zhang, Zhi-gang; Li, Wen-biao; Zhou, Xiang-fu

    2014-01-01

    Although evidence supports a role for inflammation in interstitial cystitis/bladder pain syndrome (IC/BPS), the mechanism remains unknown. We determined whether inflammation causes an elevated expression of nerve growth factor (NGF) and transient receptor potential vanilloid receptor subtype 1 (TRPV1) and correlated them with the symptoms. Bladder biopsies were obtained from 53 IC/BPS patients and 27 controls, and hematoxylin and eosin staining, immunostaining and Western blotting were performed to detect inflammation, TRPV1-immunoreactive and PGP9.5-immunoreactive nerve fibers, and NGF, respectively. Symptoms were assessed using the Pelvic Pain/Urgency/Frequency (PUF) questionnaire and pain visual analogue scale scores. Suburothelial nerve fiber density was quantified and correlated with PUF scores. Increased severity of inflammation was correlated with a higher TRPV1-immunoreactive nerve fiber density (r = 0.4113, p = 0.0024) and higher NGF levels (r = 0.3775, p = 0.0052). Suburothelial TRPV1-immunoreactive nerve fiber density was significantly correlated with pain scores and urgency scores (r = 0.3320, p = 0.0145 and r = 0.3823, p = 0.0039, respectively). PGP9.5-immunoreactive nerve fibers were significantly increased in IC/BPS (p = 0.0193) and had a positive relationship with inflammation severity (r = 0.6138, p < 0.0001). Our study revealed increased severity of inflammation correlated with a higher expression of TRPV1-immunoreactive nerve fibers and NGF in IC/BPS and correlated with clinical symptoms. 2014 S. Karger AG, Basel.

  11. Single injection of a novel nerve growth factor coacervate improves structural and functional regeneration after sciatic nerve injury in adult rats.

    PubMed

    Li, Rui; Wu, Jiang; Lin, Zhenkun; Nangle, Matthew R; Li, Yi; Cai, Pingtao; Liu, Dan; Ye, Libin; Xiao, Zecong; He, Chaochao; Ye, Jingjing; Zhang, Hongyu; Zhao, Yingzheng; Wang, Jian; Li, Xiaokun; He, Yan; Ye, Qingsong; Xiao, Jian

    2017-02-01

    The prototypical neurotrophin, nerve growth factor (NGF), plays an important role in the development and maintenance of many neurons in both the central and peripheral nervous systems, and can promote functional recovery after peripheral nerve injury in adulthood. However, repair of peripheral nerve defects is hampered by the short half-life of NGF in vivo, and treatment with either NGF alone or NGF contained in synthetic nerve conduits is inferior to the use of nerve autografts, the current gold standard. We tested the reparative ability of a single local injection of a polyvalent coacervate containing polycation-poly(ethylene argininylaspartate diglyceride; PEAD), heparin, and NGF, in adult rats following sciatic nerve crush injury, using molecular, histological and behavioral approaches. In vitro assays demonstrated that NGF was loaded into the coacervate at nearly 100% efficiency, and was protected from proteolytic degradation. In vivo, the coacervate enhanced NGF bioavailability, leading to a notable improvement in motor function (track walking analysis) after 30days. The NGF coacervate treatment was also associated with better weight gain and reduction in atrophy of the gastrocnemius muscle. Furthermore, light and electron microscopy showed that the number of myelinated axons and axon-to-fiber ratio (G-ratio) were significantly higher in NGF coacervate-treated rats compared with control groups. Expression of markers of neural tissue regeneration (MAP-2, S-100β, MBP and GAP-43), as well as proliferating Schwann cells and myelin-axon relationships (GFAP and NF200), were also increased. These observations suggest that even a single administration of NGF coacervate could have therapeutic value for peripheral nerve regeneration and functional recovery.

  12. Time-Dependent Nerve Growth Factor Signaling Changes in the Rat Retina During Optic Nerve Crush-Induced Degeneration of Retinal Ganglion Cells.

    PubMed

    Mesentier-Louro, Louise A; De Nicolò, Sara; Rosso, Pamela; De Vitis, Luigi A; Castoldi, Valerio; Leocani, Letizia; Mendez-Otero, Rosalia; Santiago, Marcelo F; Tirassa, Paola; Rama, Paolo; Lambiase, Alessandro

    2017-01-05

    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, p75(NTR), 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 p75(NTR) enhancement. In contrast, ONC-induced reduction of TrkA, and increased proNGF were observed only at 7 and 14 dac. We propose that proNGF and p75(NTR) 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.

  13. Time-Dependent Nerve Growth Factor Signaling Changes in the Rat Retina During Optic Nerve Crush-Induced Degeneration of Retinal Ganglion Cells

    PubMed Central

    Mesentier-Louro, Louise A.; De Nicolò, Sara; Rosso, Pamela; De Vitis, Luigi A.; Castoldi, Valerio; Leocani, Letizia; Mendez-Otero, Rosalia; Santiago, Marcelo F.; Tirassa, Paola; Rama, Paolo; Lambiase, Alessandro

    2017-01-01

    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. PMID:28067793

  14. Nerve growth factor promotes killing of Leishmania donovani by macrophages through the induction of hydrogen peroxide.

    PubMed

    Chiba, Rieko; Amagai, Yosuke; Tanaka, Akane; Katakura, Ken; Matsuda, Hiroshi

    2014-08-01

    Visceral leishmaniasis is protozoonosis that occurs worldwide and still requires effective therapies with less toxicity. In this study, we examined the antileishmanial effect of nerve growth factor (NGF) using a murine infection model. NGF blocked the infection of macrophages by Leishmania donovani, which was completely cancelled by a hydrogen peroxide inhibitor. In vivo, not only did NGF show antileishmanial effects, but combination therapy of NGF and sodium stibogluconate synergistically exhibited the activity more potently than each monotherapy. These results indicate that NGF exerts antileishmanial effect by stimulating hydrogen peroxide production in macrophages and can be a novel therapy for leishmaniasis.

  15. Nerve growth factor (NGF) and NGF mRNA change in rat uterus during pregnancy.

    PubMed

    Varol, F G; Duchemin, A M; Neff, N H; Hadjiconstantinou, M

    2000-11-10

    During pregnancy, the uterus undergoes a profound sympathetic denervation. To explore whether this is associated with changes in neurotrophic factors, we assayed nerve growth factor (NGF) and NGF mRNA in the uterus of non-pregnant and pregnant rats. In the uterine horn, the concentration of NGF and its mRNA decreased during middle and late pregnancy. However, when values were corrected for the increase of uterine weight and total RNA yield during pregnancy, NGF content and mRNA per horn increased during middle and late pregnancy. Similar, but less pronounced, changes were observed in the cervix. By seven days postpartum, both parameters returned to near normal.

  16. Response of the regenerating telencephalon of Lacerta viridis to nerve growth factor.

    PubMed

    Del Grande, P; Minelli, G

    1980-01-01

    To identify the nature of the dividing cells during the regenerative process of the telencephalon, the authors administered nerve growth factor (NGF) to Lacerta viridis with a wedge of telencephalon removed. Some known centers of cell proliferation were unresponsive to the treatment, whereas the ventral end of the telencephalic ventricle underwent an increase of up to 200% in proliferation rate. On the basis of this observation and data in the literature, the authors propose that the cell proliferation beginning in the medial area during the regeneration of the telencephalic ventricle is due to catecholaminergic neuroblasts still present in the adult.

  17. Altered levels of nerve growth factor in the thymus of subjects with myasthenia gravis.

    PubMed

    Stampachiacchiere, Barbara; Marinova, Tsvetana; Velikova, Kamelia; Philipov, Stanislav; Stankulov, Ivan S; Chaldakov, George N; Fiore, Marco; Aloe, Luigi

    2004-01-01

    We have previously reported that nerve growth factor (NGF), a polypeptide known for its neurotrophic activities, is also involved in the differentiation and survival of immune cells, and that NGF and its high-affinity receptor are present in the thymus. We here demonstrate that the thymus of humans affected by myasthenia gravis (MG) contains significant concentrations of NGF. These observations support our hypothesis of a role for NGF in the thymus and suggest that the changes observed in the thymus of subject with MG may have functional significance.

  18. Measuring nerve growth factor in saliva by immunoassay: A cautionary note.

    PubMed

    Matin, Marla J; Li, Daming; Peterson, Jon; Taylor, Marcus K; Laurent, Heidemarie K; Lucas, Todd; Granger, Steve J; Granger, Douglas A; Granger, Steve W

    2016-01-01

    Nerve growth factor (NGF), a neurotrophin, modulates a diverse set of physiologic processes in the nervous, immune, and endocrine systems. Studies suggest that NGF can be measured in saliva (sNGF). Historically, the method for measuring sNGF involves the off-label use of an enzyme immunoassay designed for use with cell-culture supernatants/tissue extracts (Nam et al., 2007; Ruhl et al., 2004). In a series of experiments we reveal this measurement strategy is subject to non-specific interference by constituents present in oral fluids. We conclude that the measurement of sNGF by this assay is not optimal for use with oral fluid specimens.

  19. Erythromycin and clarithromycin modulation of growth factor-induced expression of heparanase mRNA on human lung cancer cells in vitro.

    PubMed Central

    Sasaki, M; Ito, T; Kashima, M; Fukui, S; Izumiyama, N; Watanabe, A; Sano, M; Fujiwara, Y; Miura, M

    2001-01-01

    Heparanase activity is correlated with the metastatic potential of several cancer cells and is a key enzyme in the breakdown of tissue barriers. It is also involved in the regulation of growth factor and cytokine activity. However, little is known about the factors that induce heparanase in cancer cells. We investigated the effect of three growth factors, platelet-derived growth factor (PDGF), hepatocyte growth factor (HGF) and basic fibroblast growth factor (bFGF), on heparanase mRNA induction in lung cancer cells in vitro. In addition, we examined the effect of erythromycin (EM) and clarithromycin (CAM), which are 14-membered ring macrolide antibiotics that act as biological response modifiers, on the expression of heparanase mRNA induced by growth factors. PDGF, HGF and bFGF stimulated cell migration activity and enhanced the expression of heparanase mRNA in the human lung adenocarcinoma cell line A549. Via different mechanisms, EM and CAM modulate the induction by these factors of heparanase mRNA expression on A549 cells. EM also significantly suppressed A549 cell migration induced by PDGF and HGF, and CAM significantly suppressed A549cell migration induced by bFGF. The results suggest that the growth factors PDGF, HGF and bFGF are important inducers of heparanase in potentially invasive and metastatic cancer cells. The suppressive effect of heparanase mRNA expression by EM and CAM may have interestingtherapeutic applications in the prevention of metastasis. PMID:11759110

  20. The Role of Nerve Growth Factor (NGF) and Its Precursor Forms in Oral Wound Healing.

    PubMed

    Schenck, Karl; Schreurs, Olav; Hayashi, Katsuhiko; Helgeland, Kristen

    2017-02-11

    Nerve growth factor (NGF) and its different precursor forms are secreted into human saliva by salivary glands and are also produced by an array of cells in the tissues of the oral cavity. The major forms of NGF in human saliva are forms of pro-nerve growth factor (pro-NGF) and not mature NGF. The NGF receptors tropomyosin-related kinase A (TrkA) and p75 neurotrophin receptor (p75(NTR)) are widely expressed on cells in the soft tissues of the human oral cavity, including keratinocytes, endothelial cells, fibroblasts and leukocytes, and in ductal and acinar cells of all types of salivary glands. In vitro models show that NGF can contribute at most stages in the oral wound healing process: restitution, cell survival, apoptosis, cellular proliferation, inflammation, angiogenesis and tissue remodeling. NGF may therefore take part in the effective wound healing in the oral cavity that occurs with little scarring. As pro-NGF forms appear to be the major form of NGF in human saliva, efforts should be made to study its function, specifically in the process of wound healing. In addition, animal and clinical studies should be initiated to examine if topical application of pro-NGF or NGF can be a therapy for chronic oral ulcerations and wounds.

  1. Nerve growth factor-immobilized polypyrrole: Bioactive electrically conducting polymer for enhanced neurite extension

    PubMed Central

    Gomez, Natalia; Schmidt, Christine E.

    2010-01-01

    Biomaterials that present multiple stimuli are attractive for a number of biomedical applications. In particular, electrical and biological cues are important factors to include in interfaces with neurons for applications such as nerve conduits and neural probes. Here, we report the combination of these two stimuli, by immobilizing nerve growth factor (NGF) on the surface of the electrically conducting polymer polypyrrole (PPy). NGF was immobilized using an intermediate linker provided by a layer of polyallylamine conjugated to an arylazido functional group. Upon exposure to UV light and activation of the azido groups, NGF was fixed to the substrate. Three different surface concentrations were obtained (0.21–0.98 ng/mm2) and similar levels of neurite extension were observed on immobilized NGF as with soluble NGF. Additionally, electrical stimulation experiments were conducted with the modified polymer and revealed a 50% increase in neurite outgrowth in PC12 cells compared to experiments without electrical stimulation. This novel modification of PPy provides both electrical and biological stimulation, by presenting tethered growth factors and only producing a small decrease in the material's properties (conductivity ~10 S cm−1) when compared to other modification techniques (conductivity ~10−3–10−6 S cm−1. PMID:17111407

  2. Gelatin-based hydrogel for vascular endothelial growth factor release in peripheral nerve tissue engineering.

    PubMed

    Gnavi, S; di Blasio, L; Tonda-Turo, C; Mancardi, A; Primo, L; Ciardelli, G; Gambarotta, G; Geuna, S; Perroteau, I

    2017-02-01

    Hydrogels are promising materials in regenerative medicine applications, due to their hydrophilicity, biocompatibility and capacity to release drugs and growth factors in a controlled manner. In this study, biocompatible and biodegradable hydrogels based on blends of natural polymers were used in in vitro and ex vivo experiments as a tool for VEGF-controlled release to accelerate the nerve regeneration process. Among different candidates, the angiogenic factor VEGF was selected, since angiogenesis has been long recognized as an important and necessary step during tissue repair. Recent studies have pointed out that VEGF has a beneficial effect on motor neuron survival and Schwann cell vitality and proliferation. Moreover, VEGF administration can sustain and enhance the growth of regenerating peripheral nerve fibres. The hydrogel preparation process was optimized to allow functional incorporation of VEGF, while preventing its degradation and denaturation. VEGF release was quantified through ELISA assay, whereas released VEGF bioactivity was validated in human umbilical vein endothelial cells (HUVECs) and in a Schwann cell line (RT4-D6P2T) by assessing VEGFR-2 and downstream effectors Akt and Erk1/2 phosphorylation. Moreover, dorsal root ganglia explants cultured on VEGF-releasing hydrogels displayed increased neurite outgrowth, providing confirmation that released VEGF maintained its effect, as also confirmed in a tubulogenesis assay. In conclusion, a gelatin-based hydrogel system for bioactive VEGF delivery was developed and characterized for its applicability in neural tissue engineering. Copyright © 2014 John Wiley & Sons, Ltd.

  3. Postnatal expression of nerve growth factor receptors in the rat testis.

    PubMed

    Djakiew, D; Pflug, B; Dionne, C; Onoda, M

    1994-08-01

    Because nerve growth factor beta (NGF beta) and its corresponding receptors have been implicated in the paracrine regulation of spermatogenesis, we examined the postnatal developmental expression of the low- and high-affinity NGF receptors in the rat testis, and localized their expression to specific testicular cell types. The neurotropin receptors consist of a low-affinity p75 nerve growth factor receptor (LNGFR) and a family of high-affinity tyrosine receptor kinases (trk). Both the p75 LNGFR gene product and the trk receptor gene product were detected in immature rat testes, with maximal expression in 10- and 20-day-old rats. Expression of the testicular p75 LNGFR and the trk receptor progressively declined in older animals so that they were barely detectable in 90-day-old adult rats. The 75-kDa LNGFR was detected in membrane fractions of Sertoli cells, whereas the p75 LNGFR was not detected by Western blot in membrane fractions of round spermatids and primary spermatocytes. Interestingly, microsomal fractions of peritubular myoid cells were immunoreactive for a 65-kDa band on Western blots with the p75 LNGFR monoclonal antibody. Immunoblot analysis of the trk receptor in cell lysates of isolated cell types was inconclusive. Excess NGF beta and round spermatid protein, which is known to contain a NGF-like protein, were both capable of displacing the binding of 125I-NGF beta from the surface of Sertoli cells.(ABSTRACT TRUNCATED AT 250 WORDS)

  4. The Role of Nerve Growth Factor (NGF) and Its Precursor Forms in Oral Wound Healing

    PubMed Central

    Schenck, Karl; Schreurs, Olav; Hayashi, Katsuhiko; Helgeland, Kristen

    2017-01-01

    Nerve growth factor (NGF) and its different precursor forms are secreted into human saliva by salivary glands and are also produced by an array of cells in the tissues of the oral cavity. The major forms of NGF in human saliva are forms of pro-nerve growth factor (pro-NGF) and not mature NGF. The NGF receptors tropomyosin-related kinase A (TrkA) and p75 neurotrophin receptor (p75NTR) are widely expressed on cells in the soft tissues of the human oral cavity, including keratinocytes, endothelial cells, fibroblasts and leukocytes, and in ductal and acinar cells of all types of salivary glands. In vitro models show that NGF can contribute at most stages in the oral wound healing process: restitution, cell survival, apoptosis, cellular proliferation, inflammation, angiogenesis and tissue remodeling. NGF may therefore take part in the effective wound healing in the oral cavity that occurs with little scarring. As pro-NGF forms appear to be the major form of NGF in human saliva, efforts should be made to study its function, specifically in the process of wound healing. In addition, animal and clinical studies should be initiated to examine if topical application of pro-NGF or NGF can be a therapy for chronic oral ulcerations and wounds. PMID:28208669

  5. Recombinant expression of human nerve growth factor beta in rabbit bone marrow mesenchymal stem cells.

    PubMed

    Fan, Bo-Sheng; Lou, Ji-Yu

    2010-12-01

    Nerve growth factor (NGF) is required for the differentiation and maintenance of sympathetic and sensory neurons. In the present study, the recombinant expression of human nerve growth factor beta (hNGF-β) gene in rabbit bone marrow mesenchymal stem cells (rMSCs) was undertaken. Recombinant vector containing hNGF-β was constructed and transferred into rMSCs, the expressions of the exogenous in rMSCs were determined by reverse transcriptase PCR (RT-PCR), ELISA and Western blot, whereas the biological activity of recombinant hNGF-β was confirmed using PC12 cells and cultures of dorsal root ganglion neurons from chicken embryos. The results showed that the hNGF-β gene expressed successfully in the rMSCs, a polypeptide with a molecular weight of 13.2 kDa was detected. The maximal expression level of recombinant hNGF-β in rMSCs reached 126.8012 pg/10(6) cells, the mean concentration was 96.4473 pg/10(6) cells. The recombinant hNGF-β in the rMSCs showed full biological activity when compared to commercial recombinant hNGF-β.

  6. Specific binding of nerve growth factor (NGF) by murine C 1300 neuroblastoma cells.

    PubMed

    Revoltella, R; Bertolini, L; Pediconi, M; Vigneti, E

    1974-08-01

    Murine C 1300 neuroblastoma cells bind with high avidity on their membrane surface the nerve growth factor (NGF), a protein capable of inducing differentiation of sympathetic nerve cells. The total binding capacity of NGF by the cells was quantitatively measured by a radioimmunoassay technique, using (125)I-labeled NGF. An average number of about 10(6) molecules of NGF could be bound, at saturation, by each cell with an average relative association constant of about 10(7) liters/mol. Using synchronized cells, it was found, however, that either the number of molecules of ligand bound or the avidity of the binding interaction between NGF and cells varied depending upon their growth cycle, the maximal-binding occurring during the G(1) and early S phase. Binding of [(125)I]NGF was suppressed by trypsin treatment of the cells, however new receptor sites were rapidly replaced onto the membrane surface within 1-2 h. Cells exposed to 3 M KCl released into the supernate a protein product exhibiting similar high avidity for NGF. Acrylamide gel electrophoresis suggested a restricted molecular heterogeneity of this product, with a major component in the 52,000 mol wt region. Antibodies made specific to this protein were capable, in the absence of the complement, of inhibiting the binding of [(125)I]NGF by the cells and in the presence of the complement they killed them.

  7. Selective decrease in axonal nerve growth factor and insulin-like growth factor I immunoreactivity in axonopathies of unknown etiology.

    PubMed

    Fressinaud, Catherine; Jean, Isabelle; Dubas, Frédéric

    2003-05-01

    In an attempt to approach the mechanisms underlying axonopathies of unknown etiology, we have studied by immunocytochemistry the fate of several growth factors in eight of such cases that we had previously analyzed by morphometry and which were characterized by a decrease in neurofilaments and an increase in beta tubulin immunostaining. Here we establish that, contrary to beta tubulin, growth-associated protein43 (GAP-43) immunolabeling is not up-regulated in theses cases, correlating well with the failure of regeneration. Neurotrophin-3 (NT-3) and its receptor TrkC were not modified compared to controls (five cases). On the contrary, we observed in all cases a pronounced decrease in the number of fibers labeled for nerve growth factor (NGF) and insulin-like growth factor I (IGF-I), which were both approximately half of control values. This decrease could not be ascribed to the reduction in fiber density since it was also present in cases without fiber loss (isolated large fiber atrophy). The fact that only around 50% of fibers were stained, versus all fibers in controls, probably accounted for this decrease. It contrasted also with the normality of NGF and IGF-I immunolabeling in six cases of chronic inflammatory demyelinating neuropathy that were investigated in parallel. These results differ from those reported in experimental diabetic neuropathy, during which NT-3 is also decreased. A deficient supply of specific growth factors delivered by neuronal targets may be responsible for these neuropathies and their associated axonal cytoskeleton abnormalities.

  8. Nerve growth factor receptor negates the tumor suppressor p53 as a feedback regulator

    PubMed Central

    Zhou, Xiang; Hao, Qian; Liao, Peng; Luo, Shiwen; Zhang, Minhong; Hu, Guohui; Liu, Hongbing; Zhang, Yiwei; Cao, Bo; Baddoo, Melody; Flemington, Erik K; Zeng, Shelya X; Lu, Hua

    2016-01-01

    Cancer develops and progresses often by inactivating p53. Here, we unveil nerve growth factor receptor (NGFR, p75NTR or CD271) as a novel p53 inactivator. p53 activates NGFR transcription, whereas NGFR inactivates p53 by promoting its MDM2-mediated ubiquitin-dependent proteolysis and by directly binding to its central DNA binding domain and preventing its DNA-binding activity. Inversely, NGFR ablation activates p53, consequently inducing apoptosis, attenuating survival, and reducing clonogenic capability of cancer cells, as well as sensitizing human cancer cells to chemotherapeutic agents that induce p53 and suppressing mouse xenograft tumor growth. NGFR is highly expressed in human glioblastomas, and its gene is often amplified in breast cancers with wild type p53. Altogether, our results demonstrate that cancers hijack NGFR as an oncogenic inhibitor of p53. DOI: http://dx.doi.org/10.7554/eLife.15099.001 PMID:27282385

  9. A phase 1 clinical trial of nerve growth factor gene therapy for Alzheimer disease.

    PubMed

    Tuszynski, Mark H; Thal, Leon; Pay, Mary; Salmon, David P; U, Hoi Sang; Bakay, Roy; Patel, Piyush; Blesch, Armin; Vahlsing, H Lee; Ho, Gilbert; Tong, Gang; Potkin, Steven G; Fallon, James; Hansen, Lawrence; Mufson, Elliott J; Kordower, Jeffrey H; Gall, Christine; Conner, James

    2005-05-01

    Cholinergic neuron loss is a cardinal feature of Alzheimer disease. Nerve growth factor (NGF) stimulates cholinergic function, improves memory and prevents cholinergic degeneration in animal models of injury, amyloid overexpression and aging. We performed a phase 1 trial of ex vivo NGF gene delivery in eight individuals with mild Alzheimer disease, implanting autologous fibroblasts genetically modified to express human NGF into the forebrain. After mean follow-up of 22 months in six subjects, no long-term adverse effects of NGF occurred. Evaluation of the Mini-Mental Status Examination and Alzheimer Disease Assessment Scale-Cognitive subcomponent suggested improvement in the rate of cognitive decline. Serial PET scans showed significant (P < 0.05) increases in cortical 18-fluorodeoxyglucose after treatment. Brain autopsy from one subject suggested robust growth responses to NGF. Additional clinical trials of NGF for Alzheimer disease are warranted.

  10. Rita Levi-Montalcini and the discovery of NGF, the first nerve cell growth factor.

    PubMed

    Aloe, Luigi

    2011-06-01

    The nerve growth factor (NGF) is a signaling protein, discovered by Rita Levi-Montalcini in the early 1950's for its effect on growth and differentiation of specific populations of neurons of the peripheral nervous system. Originally identified as neurite outgrowth-stimulating factor, later studies revealed that the purified molecule has a number of target cells in the central nervous system and on nonneuronal cells. Moreover, recent studies showed the potential therapeutic properties of NGF in neuropathies of the central and peripheral nervous system and diseases of the eye and skin. Here I briefly describe the discovery of NGF, the early studies of Rita LeviMontalcini, a pioneer in modern neuroscience, and my scientific and human experience working in her laboratory for over 40 years.

  11. Identification of a Peripheral Nerve Neurite Growth-Promoting Activity by Development and Use of an in vitro Bioassay

    NASA Astrophysics Data System (ADS)

    Sandrock, Alfred W.; Matthew, William D.

    1987-10-01

    The effective regeneration of severed neuronal axons in the peripheral nerves of adult mammals may be explained by the presence of molecules in situ that promote the effective elongation of neurites. The absence of such molecules in the central nervous system of these animals may underlie the relative inability of axons to regenerate in this tissue after injury. In an effort to identify neurite growth-promoting molecules in tissues that support effective axonal regeneration, we have developed an in vitro bioassay that is sensitive to substrate-bound factors of peripheral nerve that influence the growth of neurites. In this assay, neonatal rat superior cervical ganglion explants are placed on longitudinal cryostat sections of fresh-frozen sciatic nerve, and the regrowing axons are visualized by catecholamine histofluorescence. Axons are found to regenerate effectively over sciatic nerve tissue sections. When ganglia are similarly explanted onto cryostat sections of adult rat central nervous system tissue, however, axonal regeneration is virtually absent. We have begun to identify the molecules in peripheral nerve that promote effective axonal regeneration by examining the effect of antibodies that interfere with the activity of previously described neurite growth-promoting factors. Axonal elongation over sciatic nerve tissue was found to be sensitive to the inhibitory effects of INO (for inhibitor of neurite outgrowth), a monoclonal antibody that recognizes and inhibits a neurite growth-promoting activity from PC-12 cell-conditioned medium. The INO antigen appears to be a molecular complex of laminin and heparan sulfate proteoglycan. In contrast, a rabbit antiserum that recognizes laminin purified from mouse Engelbreth-Holm-Swarm (EHS) sarcoma, stains the Schwann cell basal lamina of peripheral nerve, and inhibits neurite growth over purified laminin substrata has no detectable effect on the rate of axonal regeneration in our assay.

  12. Expression and significance of nerve growth factor receptor p75 in rats' cathartic colonic wall.

    PubMed

    Fan, Yi Hong; Lu, Bin; Wang, Mei; Ni, Gui Bao; Chen, Ming Tao; Xu, Yi

    2006-01-01

    To investigate the expression of nerve growth factor receptor p75 in a normal and cathartic colon and its significance in the formation of the cathartic colon in rats. Sixty Sprague-Dawley rats were equally divided into normal control group, rhubarb group and phenolphthalein group. A model of the cathartic colon was constructed by gastric infusion with rhubarb or phenolphthalein in rats. The first dose of rhubarb and phenolphthalein was both 200 mg/kg/d and was increased by 200 mg/kg/d with each passing day. The last dose of rhubarb and phenolphthalein was 3200 mg/kg/d and 4200 mg/kg/d, respectively. The transit function of colon was measured by the Chinese ink expulsion test; the p75 in colon wall was determined by the immunohistochemical method. The transit speed was much slower in the cathartic colon group than that in the control group. The imprinted Chinese ink length and the ratio of imprinted length/total colon length in the rhubarb-induced cathartic colon was significantly shorter than that of the control group (77.38 +/- 8.42 vs 94.25 +/- 7.07 cm, P < 0.01). Those in the phenolphthalein-induced group (83.38 +/- 9.75 cm) were also significantly shorter than those of the control group but to a lesser degree (P < 0.05). p75 was abundantly expressed in the submucosal nerve plexus and weakly expressed in the myenteric plexus. The expression of p75 was much higher in the rhubarb-induced group. The expression was strongly positive in the submucosal nerve plexus, significantly higher than that in the controls (P < 0.01). In the myenteric plexus, p75 was also highly expressed (P < 0.05). In the case of the phenolphthalein-induced group, the expression of p75 was positive in the submucosal nerve plexus but was positive in the myenteric plexus of three rats only. The remaining rats were negative or weakly positive. This was not significantly different from that of the control group. The abnormal expression of p75 in cathartic colon probably has some effect on the

  13. Emotional stress induced by parachute jumping enhances blood nerve growth factor levels and the distribution of nerve growth factor receptors in lymphocytes.

    PubMed

    Aloe, L; Bracci-Laudiero, L; Alleva, E; Lambiase, A; Micera, A; Tirassa, P

    1994-10-25

    We examined the plasma nerve growth factor (NGF) level and the distribution of NGF receptors in peripheral lymphocytes of young soldiers (mean age, 20-24 yr) experiencing the thrill of a novice about to make their first parachute jumps. Blood was collected from soldiers who knew they were selected to jump (n = 26), as well as from soldiers who knew they were not selected (n = 17, controls). The former group was sampled the evening before the jump and 20 min after landing. Compared with controls, NGF levels increased 84% in prejump and 107% in postjump sampling. Our studies also showed that the increase of NGF levels preceded the increase of plasma cortisol and adrenocorticotropic hormone. No changes in the baseline levels of circulating interleukin 1 beta or tumor necrosis factor were found, suggesting that the increased levels of NGF were not correlated with change in these cytokines. Moreover, immunofluorescence analysis demonstrated that parachuting stress enhances the distribution of low-affinity p75LNGFR and high-affinity p140trkA NGF receptors in circulating peripheral blood mononuclear cells. These observations suggest that the release of NGF might be involved in the activation of cells of the immune system and is most probably associated with homeostatic adaptive mechanisms, as previously shown for stressed rodents.

  14. Emotional stress induced by parachute jumping enhances blood nerve growth factor levels and the distribution of nerve growth factor receptors in lymphocytes.

    PubMed Central

    Aloe, L; Bracci-Laudiero, L; Alleva, E; Lambiase, A; Micera, A; Tirassa, P

    1994-01-01

    We examined the plasma nerve growth factor (NGF) level and the distribution of NGF receptors in peripheral lymphocytes of young soldiers (mean age, 20-24 yr) experiencing the thrill of a novice about to make their first parachute jumps. Blood was collected from soldiers who knew they were selected to jump (n = 26), as well as from soldiers who knew they were not selected (n = 17, controls). The former group was sampled the evening before the jump and 20 min after landing. Compared with controls, NGF levels increased 84% in prejump and 107% in postjump sampling. Our studies also showed that the increase of NGF levels preceded the increase of plasma cortisol and adrenocorticotropic hormone. No changes in the baseline levels of circulating interleukin 1 beta or tumor necrosis factor were found, suggesting that the increased levels of NGF were not correlated with change in these cytokines. Moreover, immunofluorescence analysis demonstrated that parachuting stress enhances the distribution of low-affinity p75LNGFR and high-affinity p140trkA NGF receptors in circulating peripheral blood mononuclear cells. These observations suggest that the release of NGF might be involved in the activation of cells of the immune system and is most probably associated with homeostatic adaptive mechanisms, as previously shown for stressed rodents. Images PMID:7937971

  15. Activity-dependent release of precursor nerve growth factor, conversion to mature nerve growth factor, and its degradation by a protease cascade.

    PubMed

    Bruno, Martin A; Cuello, A Claudio

    2006-04-25

    In this report, we provide direct demonstration that the neurotrophin nerve growth factor (NGF) is released in the extracellular space in an activity-dependent manner in its precursor form (proNGF) and that it is in this compartment that its maturation and degradation takes place because of the coordinated release and the action of proenzymes and enzyme regulators. This converting protease cascade and its endogenous regulators (including tissue plasminogen activator, plasminogen, neuroserpin, precursor matrix metalloproteinase 9, and tissue inhibitor metalloproteinase 1) are colocalized in neurons of the cerebral cortex and released upon neuronal stimulation. We also provide evidence that this mechanism operates in in vivo conditions, as the CNS application of inhibitors of converting and degrading enzymes lead to dramatic alterations in the tissue levels of either precursor NGF or mature NGF. Pathological alterations of this cascade in the CNS might cause or contribute to a lack of proper neuronal trophic support in conditions such as cerebral ischemia, seizure and Alzheimer's disease or, conversely, to excessive local production of neurotrophins as reported in inflammatory arthritis pain.

  16. Activity-dependent release of precursor nerve growth factor, conversion to mature nerve growth factor, and its degradation by a protease cascade

    PubMed Central

    Bruno, Martin A.; Cuello, A. Claudio

    2006-01-01

    In this report, we provide direct demonstration that the neurotrophin nerve growth factor (NGF) is released in the extracellular space in an activity-dependent manner in its precursor form (proNGF) and that it is in this compartment that its maturation and degradation takes place because of the coordinated release and the action of proenzymes and enzyme regulators. This converting protease cascade and its endogenous regulators (including tissue plasminogen activator, plasminogen, neuroserpin, precursor matrix metalloproteinase 9, and tissue inhibitor metalloproteinase 1) are colocalized in neurons of the cerebral cortex and released upon neuronal stimulation. We also provide evidence that this mechanism operates in in vivo conditions, as the CNS application of inhibitors of converting and degrading enzymes lead to dramatic alterations in the tissue levels of either precursor NGF or mature NGF. Pathological alterations of this cascade in the CNS might cause or contribute to a lack of proper neuronal trophic support in conditions such as cerebral ischemia, seizure and Alzheimer’s disease or, conversely, to excessive local production of neurotrophins as reported in inflammatory arthritis pain. PMID:16618925

  17. Glycogen Synthase Kinase 3β Is a Negative Regulator of Growth Factor-induced Activation of the c-Jun N-terminal Kinase*

    PubMed Central

    Liu, Shuying; Yu, Shuangxing; Hasegawa, Yutaka; LaPushin, Ruth; Xu, Hong-Ji; Woodgett, James R.; Mills, Gordon B.; Fang, Xianjun

    2016-01-01

    The c-Jun N-terminal kinase (JNK)/stress activated protein kinase is preferentially activated by stress stimuli. Growth factors, particularly ligands for G protein-coupled receptors, usually induce only modest JNK activation, although they may trigger marked activation of the related extracellular signal-regulated kinase. In the present study, we demonstrated that homozygous disruption of glycogen synthase kinase 3β (GSK-3β) dramatically sensitized mouse embryonic fibroblasts (MEFs) to JNK activation induced by lysophosphatidic acid (LPA) and sphingosine-1-phosphate, two prototype ligands for G protein-coupled receptors. To a lesser degree, a lack of GSK-3β also potentiated JNK activation in response to epidermal growth factor. In contrast, the absence of GSK-3β decreased UV light-induced JNK activation. The increased JNK activation induced by LPA in GSK-3β null MEFs was insufficient to trigger apoptotic cell death or growth inhibition. Instead, the increased JNK activation observed in GSK-3β−/− MEFs was associated with an increased proliferative response to LPA, which was reduced by the inhibition of JNK. Ectopic expression of GSK-3β in GSK-3β-negative MEFs restrained LPA-triggered JNK phosphorylation and induced a concomitant decrease in the mitogenic response to LPA compatible with GSK-3β through the inhibition of JNK activation, thus limiting LPA-induced cell proliferation. Mutation analysis indicated that GSK-3β kinase activity was required for GSK-3β to optimally inhibit LPA-stimulated JNK activation. Thus GSK-3β serves as a physiological switch to specifically repress JNK activation in response to LPA, sphingosine-1-phosphate, or the epidermal growth factor. These results reveal a novel role for GSK-3β in signal transduction and cellular responses to growth factors. PMID:15466414

  18. PC12 cell mutants that possess low- but not high-affinity nerve growth factor receptors neither respond to nor internalize nerve growth factor

    PubMed Central

    1986-01-01

    Four mutant PC12 pheochromocytoma cell lines that are nerve growth factor (NGF)-nonresponsive (PC12nnr) have been selected from chemically mutagenized cultures by a double selection procedure: failure both to grow neurites in the presence of NGF and to survive in NGF-supplemented serum-free medium. The PC12nnr cells were deficient in all additional NGF responses surveyed: abatement of cell proliferation, changes in glycoprotein composition, induction of ornithine decarboxylase, rapid changes in protein phosphorylation, and cell surface ruffling. However, PC12nnr cells closely resembled non-NGF-treated PC12 cells in most properties tested: cell size and shape; division rate; protein, phosphoprotein, and glycoprotein composition; and cell surface morphology. All four PC12nnr lines differed from PC12 cells in three ways in addition to failure of NGF response: PC12nnr cells failed to internalize bound NGF by the normal, saturable, high-affinity mechanism present in PC12 cells. The PC12nnr cells bound NGF but entirely, or nearly entirely, at low-affinity sites only, whereas PC12 cells possess both high- and low-affinity NGF binding sites. The responses to dibutyryl cyclic AMP that were tested appeared to be enhanced or altered in the PC12nnr cells compared to PC12 cells. Internalization of, and responses to, epidermal growth factor were normal in the PC12nnr cells ruling out a generalized defect in hormonal binding, uptake, or response mechanisms. These findings are consistent with a causal association between the presence of high-affinity NGF receptors and of NGF responsiveness and internalization. A possible relationship is also suggested between regulation of cAMP responses and regulation of NGF responses or NGF receptor affinity. PMID:3005338

  19. Nerve growth factor injected systemically improves the recovery of the inferior alveolar nerve in a rabbit model of mandibular distraction osteogenesis.

    PubMed

    Du, Zhao-jie; Wang, Lei; Lei, De-lin; Liu, Bao-lin; Cao, Jian; Zhang, Pu; Ma, Qin

    2011-10-01

    Our aim was to find out if nerve growth factor (NGF) injected systemically could improve the recovery of the inferior alveolar nerve in a rabbit model of mandibular distraction osteogenesis. We used 48 New Zealand white rabbits that were treated with bilateral distraction osteogenesis at a rate of 0.5mm/12h for 10 days. Immediately postoperatively, NGF or sodium chloride 0.6 μg/day was injected intramuscularly for 20 days. At the end of distraction and after consolidation times of 1, 2, and 4 weeks, the inferior alveolar nerves were evaluated histologically and histomorphometrically. Histologically, at 2 and 4 weeks there was less myelin debris, and more regenerating axons were present, in the NGF than the control groups. The density of myelinated axons was significantly greater in groups with NGF than controls at 2 and 4 weeks (p<0.05). NGF given systemically can accelerate the recovery of the inferior alveolar nerve in rabbits after mandibular distraction osteogenesis, and is a promising treatment option for neurological complications of mandibular distraction osteogenesis.

  20. Grb2 negatively regulates epidermal growth factor-induced phospholipase C-gamma1 activity through the direct interaction with tyrosine-phosphorylated phospholipase C-gamma1.

    PubMed

    Choi, Jang Hyun; Hong, Won-Pyo; Yun, Sanguk; Kim, Hyeon Soo; Lee, Jong-Ryul; Park, Jong Bae; Bae, Yun Soo; Ryu, Sung Ho; Suh, Pann-Ghill

    2005-10-01

    Phospholipase C-gamma1 (PLC-gamma1) plays pivotal roles in cellular growth and proliferation. Upon the stimulation of growth factors and hormones, PLC-gamma1 is rapidly phosphorylated at three known sites; Tyr771, Tyr783 and Tyr1254 and its enzymatic activity is up-regulated. In this study, we demonstrate for the first time that Grb2, an adaptor protein, specifically interacts with tyrosine-phosphorylated PLC-gamma1 at Tyr783. The association of Grb2 with PLC-gamma1 was induced by the treatment with epidermal growth factor (EGF). Replacement of Tyr783 with Phe completely blocked EGF-induced interaction of PLC-gamma1 with Grb2, indicating that tyrosine phosphorylation of PLC-gamma1 at Tyr783 is essential for the interaction with Grb2. Interestingly, the depletion of Grb2 from HEK-293 cells by RNA interference significantly enhanced increased EGF-induced PLC-gamma1 enzymatic activity and mobilization of the intracellular Ca2+, while it did not affect EGF-induced tyrosine phosphorylation of PLC-gamma1. Furthermore, overexpression of Grb2 inhibited PLC-gamma1 enzymatic activity. Taken together, these results suggest Grb2, in addition to its key function in signaling through Ras, may have a negatively regulatory role on EGF-induced PLC-gamma1 activation.

  1. Vascular endothelial growth factor promotes anatomical and functional recovery of injured peripheral nerves in the avascular cornea.

    PubMed

    Pan, Zan; Fukuoka, Shima; Karagianni, Natalia; Guaiquil, Victor H; Rosenblatt, Mark I

    2013-07-01

    Peripheral nerve injury is a major neurological disorder that can cause severe motor and sensory dysfunction. Neurogenic effects of vascular endothelial growth factor (VEGF) have been found in the central nervous system, and we examined whether VEGF could promote anatomical and functional recovery of peripheral nerves after injury using an avascular corneal nerve injury model. We found that VEGF enhanced neurite elongation in isolated trigeminal ganglion neurons in a dose-dependent manner. This effect was suppressed by neutralizing antibodies for VEGF receptor (VEGFR) 1 and 2 or neuropilin receptor 1 or by VEGFR2 inhibitors (SU 1498 and Ki 8751). In vivo, mice receiving sustained VEGF via implanted pellets showed increased corneal nerve regeneration after superficial injury compared with those receiving vehicle. VEGF injected subconjunctivally at the time of injury accelerated reinnervation, the recovery of mechanosensation, and epithelial wound healing. Endogenous VEGF expression was up-regulated in the corneal epithelium and stroma after wounding. Thus, VEGF can mediate peripheral neuron growth but requires the activation of multiple VEGF receptor types. In addition, VEGF can accelerate the return of sensory and trophic functions of damaged peripheral nerves. Wounding induces the expression of VEFG, which may modulate physiological nerve repair.

  2. Schwann cell-neuronal interactions in the rat involve nerve growth factor.

    PubMed

    Urschel, B A; Hulsebosch, C E

    1990-06-01

    To gain some insight into possible functions of nerve growth factor (NGF), we suppressed the endogenous levels of NGF in newborn rats by subcutaneous injections (3 microliters/g body weight) of rabbit antibodies to purified mouse beta-NGF (ANTI-NGF). Fiber and axonal areas and perimeters were measured for unmyelinated and myelinated sensory fibers in T9 dorsal roots (DR) in three groups of animals: 1) ANTI-NGF treated littermates, 2) preimmune sera treated littermates (PREIMM), and 3) untreated littermates (UNTR). In some rats, fibers in ventral roots (VR) were measured and, in other rats, sensory processes in peripheral nerves (PN) were measured following radical ventral rhizotomy. The only outer area and perimeter measurements that were statistically different were those in the ventral root (P less than 0.013 and P less than 0.043, respectively). However, myelin thickness was significantly thinner in the dorsal roots of the ANTI-NGF group than in the dorsal roots of the UNTR and PREIMM groups (P less than 0.000009 and P less than 10(-6), respectively). Myelin thickness in the ventral roots of the ANTI-NGF group was also statistically thinner than that in the UNTR group (P less than 0.001). There were no statistically significant differences when comparing the UNTR group to the PREIMM group. In the peripheral nerves studied, there was no significant change in the myelin thickness between the ANTI-NGF and UNTR groups of animals. These results indicate that Schwann cell-neuronal interactions are altered by the inactivation of NGF, and that 1) the central processes of sensory fibers are affected and not the peripheral processes and 2) motor fiber myelination is altered.(ABSTRACT TRUNCATED AT 250 WORDS)

  3. [Effect of embryonic anlage allografts of the rat spinal cord on growth of regenerating fibers of the recipient nerve].

    PubMed

    Petrova, E S; Isaeva, E N

    2014-01-01

    A comparative study of the effect of tissue and suspension allografts of an embryonic spinal cord on regeneration of nerve fibers of impaired (by application of a ligature) sciatic nerve in rats was conducted. It was demonstrated that unlike tissue grafts that reach a large volume 21 and 60 days after transplantation, suspension grafts do not inhibit the growth of axons of the recipient to the periphery. It was established that introduction of a suspension of dissociated cells of the spinal cord embryonic anlages (but not fragments of these anlages) into the impaired sciatic nerve in rats results in an increase in the amount of myelinated regenerating nerve fibers of the recipient 60 days after the operation.

  4. Superoxide dismutase abolishes the platelet-derived growth factor-induced release of prostaglandin E2 by blocking induction of nitric oxide synthase: role of superoxide.

    PubMed

    Kelner, M J; Uglik, S F

    1995-09-10

    The ability of platelet-derived growth factor (PDGF) to induce prostaglandin E2 (PGE2) release in fibroblasts is abolished when copper-zinc superoxide dismutase activity is increased by transfection of an expression vector. The effect is specific to copper-zinc superoxide dismutase as glutathione peroxidase-overexpressing NIH3T3 cells, again produced by transfection of an expression vector, retain the ability to release PGE2 in response to growth factor stimulation. The defect in PDGF-induced PGE2 release occurs prior to action of prostaglandin H synthase/cyclooxygenase as release of arachadonic acid (in response to PDGF) does not occur in the superoxide dismutase-overexpressing clones. The defect in PDGF-induced release of PGE2 in superoxide dismutase-overexpressing clones differs from the defect found in pEJ-ras-transformed clones. The parent cells, the glutathione peroxidase-expressing cells, and the superoxide dismutase-overexpressing cells all release PGE2 in response to exogenous nitric oxide, whereas the pEJ-ras-transformed cells do not. The glutathione peroxidase-expressing cells also retained the ability to release nitrite in response to PDGF, whereas the superoxide dismutase-expressing clones do not. PDGF stimulates nitric oxide synthase activity in NIH3T3 cells, but not in the superoxide dismutase-expressing clones. These results indicate that superoxide dismutase overexpression blocks the PDGF-induced release of PGE2 by blocking induction of nitric oxide synthase. This indicates that the increase of nitric oxide synthase induced by PDGF is mediated in part by production of superoxide. These findings link cellular oxygen radical homeostasis to three different classes of messenger molecules (growth factors, nitric oxide, and prostaglandins).

  5. The statistical mechanics of complex signaling networks: nerve growth factor signaling

    NASA Astrophysics Data System (ADS)

    Brown, K. S.; Hill, C. C.; Calero, G. A.; Myers, C. R.; Lee, K. H.; Sethna, J. P.; Cerione, R. A.

    2004-10-01

    The inherent complexity of cellular signaling networks and their importance to a wide range of cellular functions necessitates the development of modeling methods that can be applied toward making predictions and highlighting the appropriate experiments to test our understanding of how these systems are designed and function. We use methods of statistical mechanics to extract useful predictions for complex cellular signaling networks. A key difficulty with signaling models is that, while significant effort is being made to experimentally measure the rate constants for individual steps in these networks, many of the parameters required to describe their behavior remain unknown or at best represent estimates. To establish the usefulness of our approach, we have applied our methods toward modeling the nerve growth factor (NGF)-induced differentiation of neuronal cells. In particular, we study the actions of NGF and mitogenic epidermal growth factor (EGF) in rat pheochromocytoma (PC12) cells. Through a network of intermediate signaling proteins, each of these growth factors stimulates extracellular regulated kinase (Erk) phosphorylation with distinct dynamical profiles. Using our modeling approach, we are able to predict the influence of specific signaling modules in determining the integrated cellular response to the two growth factors. Our methods also raise some interesting insights into the design and possible evolution of cellular systems, highlighting an inherent property of these systems that we call 'sloppiness.'

  6. BAG-1 enhances cell-cell adhesion, reduces proliferation and induces chaperone-independent suppression of hepatocyte growth factor-induced epidermal keratinocyte migration

    SciTech Connect

    Hinitt, C.A.M.; Wood, J.; Lee, S.S.; Williams, A.C.; Howarth, J.L.; Glover, C.P.; Uney, J.B.; Hague, A.

    2010-08-01

    Cell motility is important in maintaining tissue homeostasis, facilitating epithelial wound repair and in tumour formation and progression. The aim of this study was to determine whether BAG-1 isoforms regulate epidermal cell migration in in vitro models of wound healing. In the human epidermal cell line HaCaT, endogenous BAG-1 is primarily nuclear and increases with confluence. Both transient and stable p36-Bag-1 overexpression resulted in increased cellular cohesion. Stable transfection of either of the three human BAG-1 isoforms p36-Bag-1 (BAG-1S), p46-Bag-1 (BAG-1M) and p50-Bag-1 (BAG-1L) inhibited growth and wound closure in serum-containing medium. However, in response to hepatocyte growth factor (HGF) in serum-free medium, BAG-1S/M reduced communal motility and colony scattering, but BAG-1L did not. In the presence of HGF, p36-Bag-1 transfectants retained proliferative response to HGF with no change in ERK1/2 activation. However, the cells retained E-cadherin localisation at cell-cell junctions and exhibited pronounced cortical actin. Point mutations in the BAG domain showed that BAG-1 inhibition of motility is independent of its function as a chaperone regulator. These findings are the first to suggest that BAG-1 plays a role in regulating cell-cell adhesion and suggest an important function in epidermal cohesion.

  7. Sulindac sulfide inhibits epidermal growth factor-induced phosphorylation of extracellular-regulated kinase 1/2 and Bad in human colon cancer cells.

    PubMed

    Rice, Pamela L; Washington, Michele; Schleman, Shea; Beard, K Scott; Driggers, Linda J; Ahnen, Dennis J

    2003-02-01

    Colorectal cancer is the second leading cause of cancer death in the United States. Nonsteroidal anti-inflammatory drugs including sulindac are promising chemopreventive agents for colorectal cancer. Sulindac and selective cyclooxygenase (COX)-2 inhibitors cause regression of colonic polyps in familial polyposis patients. Sulindac induces apoptotic cell death in cancer cells in vitro and in vivo. In tumor cells, activation of extracellular-regulated kinase (ERK) 1/2 results in phosphorylation of several ERK1/2 effectors, including the proapoptotic protein Bad. Phosphorylation of Ser112 by ERK1/2 inactivates Bad and protects the tumor cell from apoptosis. Sulindac metabolites and other nonsteroidal anti-inflammatory drugs selectively inhibit ERK1/2 phosphorylation in human colon cancer cells. In this study we show that epidermal growth factor (EGF) strongly induces phosphorylation of ERK1/2 and Bad in HT29 colon cancer cells. EGF-stimulated phosphorylation of ERK and Bad is blocked by pretreatment with U0126, a selective MAP kinase kinase (MKK)1/2 inhibitor. Similarly, pretreatment with sulindac sulfide blocks the ability of EGF to induce ERK1/2 and Bad phosphorylation, but also down-regulates total Bad but not ERK1/2 protein levels. The ability of sulindac to block ERK1/2 signaling by the EGF receptor may account for at least part of its potent growth-inhibitory effects against cancer cells.

  8. Intracellular pH regulation by Na⁺/H⁺ exchanger-1 (NHE1) is required for growth factor-induced mammary branching morphogenesis.

    PubMed

    Jenkins, Edmund C; Debnath, Shawon; Gundry, Stephen; Gundry, Sajini; Uyar, Umit; Fata, Jimmie E

    2012-05-01

    Regulation of intracellular pH (pHi) and protection against cytosolic acidification is primarily a function of the ubiquitous plasma membrane Na+/H+exchanger-1 (NHE1), which uses a highly conserved process to transfer cytosolic hydrogen ions (H+) across plasma membranes in exchange for extracellular sodium ions (Na+). Growth factors, which are essential regulators of morphogenesis, have also been found to be key activators of NHE1 exchanger activity; however, the crosstalk between both has not been fully evaluated during organ development. Here we report that mammary branching morphogenesis induced by transforming growth factor-alpha (TGFα) requires PI3K-dependent NHE1-activation and subsequent pHi alkalization. Inhibiting NHE1 activity after TGFα stimulation with 10 μM of the NHE1-specific inhibitor N-Methyl-N-isobutyl Amiloride (MIA) dramatically disrupted branching morphogenesis, induced extensive proliferation, ectopic expression of the epithelial hyper-proliferative marker Keratin-6 and sustained activation of MAPK. Together these findings indicate a novel developmental signaling cascade involving TGFα>PI3K>NHE1>pHi alkalization, which leads to a permissible environment for MAPK negative feedback inhibition and thus regulated mammary branching morphogenesis.

  9. Real-time imaging elucidates the role of H2O2 in regulating kinetics of epidermal growth factor-induced and Src-mediated tyrosine phosphorylation signaling

    NASA Astrophysics Data System (ADS)

    Su, Ting; Li, Xiangyong; Liu, Nisha; Pan, Shaotao; Lu, Jinling; Yang, Jie; Zhang, Zhihong

    2012-07-01

    Reversible oxidation is emerging as an important regulatory mechanism in protein tyrosine phosphorylation. Generation of hydrogen peroxide (H2O2), upon growth factor stimulation, is hypothesized to inhibit activity of protein tyrosine phosphatases (PTPs). This ensures that protein tyrosine kinases can elevate the steady-state level of protein tyrosine phosphorylation, which then allows propagation of the tyrosine phosphorylation signal. However, the effects of H2O2 on the kinetics of tyrosine phosphorylation signaling remain poorly understood, especially in living cells. Therefore, we used a genetically encoded Src kinase-specific biosensor based on fluorescence resonance energy transfer (FRET) to image the kinetics of the Src-mediated tyrosine phosphorylation signaling (Src signaling) induced by epidermal growth factor (EGF). We examined the kinetics under increased and decreased H2O2 levels. Through a straightforward, quantitative analysis method which characterized the signaling kinetics, we demonstrated that H2O2 modulated the amplitude and duration of the signal by inhibiting PTPs' activity. Our evidence also suggested the effect of H2O2 on Src activation is mediated by H2O2-dependent inhibition of PTPs. Furthermore, we provide evidence showing global elevation of intracellular H2O2 level attenuates EGF-induced Src signaling.

  10. The small GTPase ROP10 of Medicago truncatula is required for both tip growth of root hairs and nod factor-induced root hair deformation.

    PubMed

    Lei, Ming-Juan; Wang, Qi; Li, Xiaolin; Chen, Aimin; Luo, Li; Xie, Yajun; Li, Guan; Luo, Da; Mysore, Kirankumar S; Wen, Jiangqi; Xie, Zhi-Ping; Staehelin, Christian; Wang, Yan-Zhang

    2015-03-01

    Rhizobia preferentially enter legume root hairs via infection threads, after which root hairs undergo tip swelling, branching, and curling. However, the mechanisms underlying such root hair deformation are poorly understood. Here, we showed that a type II small GTPase, ROP10, of Medicago truncatula is localized at the plasma membrane (PM) of root hair tips to regulate root hair tip growth. Overexpression of ROP10 and a constitutively active mutant (ROP10CA) generated depolarized growth of root hairs, whereas a dominant negative mutant (ROP10DN) inhibited root hair elongation. Inoculated with Sinorhizobium meliloti, the depolarized swollen and ballooning root hairs exhibited extensive root hair deformation and aberrant infection symptoms. Upon treatment with rhizobia-secreted nodulation factors (NFs), ROP10 was transiently upregulated in root hairs, and ROP10 fused to green fluorescent protein was ectopically localized at the PM of NF-induced outgrowths and curls around rhizobia. ROP10 interacted with the kinase domain of the NF receptor NFP in a GTP-dependent manner. Moreover, NF-induced expression of the early nodulin gene ENOD11 was enhanced by the overexpression of ROP10 and ROP10CA. These data suggest that NFs spatiotemporally regulate ROP10 localization and activity at the PM of root hair tips and that interactions between ROP10 and NF receptors are required for root hair deformation and continuous curling during rhizobial infection.

  11. Balanced levels of nerve growth factor are required for normal pregnancy progression.

    PubMed

    Frank, Pierre; Barrientos, Gabriela; Tirado-González, Irene; Cohen, Marie; Moschansky, Petra; Peters, Eva M; Klapp, Burghard F; Rose, Matthias; Tometten, Mareike; Blois, Sandra M

    2014-08-01

    Nerve growth factor (NGF), the first identified member of the family of neurotrophins, is thought to play a critical role in the initiation of the decidual response in stress-challenged pregnant mice. However, the contribution of this pathway to physiological events during the establishment and maintenance of pregnancy remains largely elusive. Using NGF depletion and supplementation strategies alternatively, in this study, we demonstrated that a successful pregnancy is sensitive to disturbances in NGF levels in mice. Treatment with NGF further boosted fetal loss rates in the high-abortion rate CBA/J x DBA/2J mouse model by amplifying a local inflammatory response through recruitment of NGF-expressing immune cells, increased decidual innervation with substance P(+) nerve fibres and a Th1 cytokine shift. Similarly, treatment with a NGF-neutralising antibody in BALB/c-mated CBA/J mice, a normal-pregnancy model, also induced abortions associated with increased infiltration of tropomyosin kinase receptor A-expressing NK cells to the decidua. Importantly, in neither of the models, pregnancy loss was associated with defective ovarian function, angiogenesis or placental development. We further demonstrated that spontaneous abortion in humans is associated with up-regulated synthesis and an aberrant distribution of NGF in placental tissue. Thus, a local threshold of NGF expression seems to be necessary to ensure maternal tolerance in healthy pregnancies, but when surpassed may result in fetal rejection due to exacerbated inflammation.

  12. Nerve growth factor-immobilized electrically conducting fibrous scaffolds for potential use in neural engineering applications.

    PubMed

    Lee, Jae Y; Bashur, Chris A; Milroy, Craig A; Forciniti, Leandro; Goldstein, Aaron S; Schmidt, Christine E

    2012-03-01

    Engineered scaffolds simultaneously exhibiting multiple cues are highly desirable for neural tissue regeneration. To this end, we developed a neural tissue engineering scaffold that displays submicrometer-scale features, electrical conductivity, and neurotrophic activity. Specifically, electrospun poly(lactic acid-co-glycolic acid) (PLGA) nanofibers were layered with a nanometer thick coating of electrically conducting polypyrrole (PPy) presenting carboxylic groups. Then, nerve growth factor (NGF) was chemically immobilized onto the surface of the fibers. These NGF-immobilized PPy-coated PLGA (NGF-PPyPLGA) fibers supported PC12 neurite formation ( 28.0±3.0% of the cells) and neurite outgrowth (14.2 μm median length), which were comparable to that observed with NGF (50 ng/mL) in culture medium ( 29.0±1.3%, 14.4 μm). Electrical stimulation of PC12 cells on NGF-immobilized PPyPLGA fiber scaffolds was found to further improve neurite development and neurite length by 18% and 17%, respectively, compared to unstimulated cells on the NGF-immobilized fibers. Hence, submicrometer-scale fibrous scaffolds that incorporate neurotrophic and electroconducting activities may serve as promising neural tissue engineering scaffolds such as nerve guidance conduits.

  13. Nerve Growth Factor-Immobilized Electrically Conducting Fibrous Scaffolds for Potential Use in Neural Engineering Applications

    PubMed Central

    Lee, Jae Y.; Bashur, Chris A.; Milroy, Craig A.; Forciniti, Leandro; Goldstein, Aaron S.

    2015-01-01

    Engineered scaffolds simultaneously exhibiting multiple cues are highly desirable for neural tissue regeneration. To this end, we developed a neural tissue engineering scaffold that displays submicrometer-scale features, electrical conductivity, and neurotrophic activity. Specifically, electrospun poly(lactic acid-co-glycolic acid) (PLGA) nanofibers were layered with a nanometer thick coating of electrically conducting polypyrrole (PPy) presenting carboxylic groups. Then, nerve growth factor (NGF) was chemically immobilized onto the surface of the fibers. These NGF-immobilized PPy-coated PLGA (NGF-PPyPLGA) fibers supported PC12 neurite formation (28.0±3.0% of the cells) and neurite outgrowth (14.2 µm median length), which were comparable to that observed with NGF (50 ng/mL) in culture medium (29.0±1.3%, 14.4 µm). Electrical stimulation of PC12 cells on NGF-immobilized PPyPLGA fiber scaffolds was found to further improve neurite development and neurite length by 18% and 17%, respectively, compared to unstimulated cells on the NGF-immobilized fibers. Hence, submicrometer-scale fibrous scaffolds that incorporate neurotrophic and electroconducting activities may serve as promising neural tissue engineering scaffolds such as nerve guidance conduits. PMID:21712166

  14. Pincher, a pinocytic chaperone for nerve growth factor/TrkA signaling endosomes

    PubMed Central

    Shao, Yufang; Akmentin, Wendy; Toledo-Aral, Juan Jose; Rosenbaum, Julie; Valdez, Gregorio; Cabot, John B.; Hilbush, Brian S.; Halegoua, Simon

    2002-01-01

    Acentral tenet of nerve growth factor (NGF) action that is poorly understood is its ability to mediate cytoplasmic signaling, through its receptor TrkA, that is initiated at the nerve terminal and conveyed to the soma. We identified an NGF-induced protein that we termed Pincher (pinocytic chaperone) that mediates endocytosis and trafficking of NGF and its receptor TrkA. In PC12 cells, overexpression of Pincher dramatically stimulated NGF-induced endocytosis of TrkA, unexpectedly at sites of clathrin-independent macropinocytosis within cell surface ruffles. Subsequently, a system of Pincher-containing tubules mediated the delivery of NGF/TrkA-containing vesicles to cytoplasmic accumulations. These vesicles selectively and persistently mediated TrkA-erk5 mitogen-activated protein kinase signaling. A dominant inhibitory mutant form of Pincher inhibited the NGF-induced endocytosis of TrkA, and selectively blocked TrkA-mediated cytoplasmic signaling of erk5, but not erk1/2, kinases. Our results indicate that Pincher mediates pinocytic endocytosis of functionally specialized NGF/TrkA endosomes with persistent signaling potential. PMID:12011113

  15. A Cell Line Producing Recombinant Nerve Growth Factor Evokes Growth Responses in Intrinsic and Grafted Central Cholinergic Neurons

    NASA Astrophysics Data System (ADS)

    Ernfors, Patrik; Ebendal, Ted; Olson, Lars; Mouton, Peter; Stromberg, Ingrid; Persson, Hakan

    1989-06-01

    The rat β nerve growth factor (NGF) gene was inserted into a mammalian expression vector and cotransfected with a plasmid conferring resistance to neomycin into mouse 3T3 fibroblasts. From this transfection a stable cell line was selected that contains several hundred copies of the rat NGF gene and produces excess levels of recombinant NGF. Such genetically modified cells were implanted into the rat brain as a probe for in vivo effects of NGF on central nervous system neurons. In a model of the cortical cholinergic deficits in Alzheimer disease, we demonstrate a marked increase in the survival of, and fiber outgrowth from, grafts of fetal basal forebrain cholinergic neurons, as well as stimulation of fiber formation by intact adult intrinsic cholinergic circuits in the cerebral cortex. Adult cholinergic interneurons in intact striatum also sprout vigorously toward implanted fibroblasts. Our results suggest that this model has implications for future treatment of neurodegenerative diseases.

  16. Ellagitannin-rich cloudberry inhibits hepatocyte growth factor induced cell migration and phosphatidylinositol 3-kinase/AKT activation in colon carcinoma cells and tumors in Min mice

    PubMed Central

    Pajari, Anne-Maria; Päivärinta, Essi; Paavolainen, Lassi; Vaara, Elina; Koivumäki, Tuuli; Garg, Ritu; Heiman-Lindh, Anu; Mutanen, Marja; Marjomäki, Varpu; Ridley, Anne J.

    2016-01-01

    Berries have been found to inhibit colon carcinogenesis in animal models, and thus represent a potential source of compounds for prevention and treatment of colorectal cancer. The mechanistic basis for their effects is not well understood. We used human colon carcinoma cells and Min mice to investigate the effects of ellagitannin-rich cloudberry (Rubus chamaemorus) extract on cancer cell migration and underlying cell signaling. Intrinsic and hepatocyte growth factor (HGF) -induced cell motility in human HT29 and HCA7 colon carcinoma cells was assessed carrying out cell scattering and scratch wound healing assays using time-lapse microscopy. Activation of Met, AKT, and ERK in cell lines and tumors of cloudberry-fed Min mice were determined using immunoprecipitation, Western blot and immunohistochemical analyses. Cloudberry extract significantly inhibited particularly HGF-induced cancer cell migration in both cell lines. Cloudberry extract inhibited the Met receptor tyrosine phosphorylation by HGF and strongly suppressed HGF-induced AKT and ERK activation in both HT29 and HCA7 cells. Consistently, cloudberry feeding (10% w/w freeze-dried berries in diet for 10 weeks) reduced the level of active AKT and prevented phosphoMet localization at the edges in tumors of Min mice. These results indicate that cloudberry reduces tumor growth and cancer cell motility by inhibiting Met signaling and consequent activation of phosphatidylinositol 3-kinase/AKT in vitro and in tumors in vivo. As the Met receptor is recognized to be a major target in cancer treatment, our results suggest that dietary phytochemicals may have therapeutic value in reducing cancer progression and metastasis. PMID:27270323

  17. Epidermal growth factor-induced proliferation of collecting duct cells from Oak Ridge polycystic kidney mice involves activation of Na+/H+ exchanger.

    PubMed

    Coaxum, Sonya D; Blanton, Mary G; Joyner, Alisha; Akter, Tanjina; Bell, P Darwin; Luttrell, Louis M; Raymond, John R; Lee, Mi-Hye; Blichmann, Paul A; Garnovskaya, Maria N; Saigusa, Takamitsu

    2014-09-15

    Epidermal growth factor (EGF) is linked to the pathogenesis of polycystic kidney disease (PKD). We explored signaling pathways activated by EGF in orpk cilia (-) collecting duct cell line derived from a mouse model of PKD (hypomorph of the Tg737/Ift88 gene) with severely stunted cilia, and in a control orpk cilia (+) cell line with normal cilia. RT-PCR demonstrated mRNAs for EGF receptor subunits ErbB1, ErbB2, ErbB3, ErbB4, and mRNAs for Na(+)/H(+) exchangers (NHE), NHE-1, NHE-2, NHE-3, NHE-4, and NHE-5 in both cell lines. EGF stimulated proton efflux in both cell lines. This effect was significantly attenuated by MIA, 5-(n-methyl-N-isobutyl) amiloride, a selective inhibitor of NHE-1 and NHE-2, and orpk cilia (-) cells were more sensitive to MIA than control cells (P < 0.01). EGF significantly induced extracellular signal-regulated kinase (ERK) phosphorylation in both cilia (+) and cilia (-) cells (63.3 and 123.6%, respectively), but the effect was more pronounced in orpk cilia (-) cells (P < 0.01). MIA significantly attenuated EGF-induced ERK phosphorylation only in orpk cilia (-) cells (P < 0.01). EGF increased proliferation of orpk cilia (+) cells and orpk cilia (-) cells, respectively, and MIA at 1-5 μM attenuated EGF-induced proliferation in orpk cilia (-) cells without affecting proliferation of orpk cilia (+) cells. EGF-induced proliferation of both cell lines was significantly decreased by the EGFR tyrosine kinase inhibitor AG1478 and MEK inhibitor PD98059. These results suggest that EGF exerts mitogenic effects in the orpk cilia (-) cells via activation of growth-associated amiloride-sensitive NHEs and ERK.

  18. Growth-associated protein 43 in differentiating peripheral nerve sheath tumors from other non-neural spindle cell neoplasms.

    PubMed

    Chen, Wei-Shen; Chen, Pei-Ling; Lu, Dongsi; Lind, Anne C; Dehner, Louis P

    2014-02-01

    The malignant peripheral nerve sheath tumor is a relatively uncommon type of soft tissue sarcoma arising from a peripheral nerve or extraneural soft tissues and showing nerve sheath differentiation. The diagnosis of malignant peripheral nerve sheath tumor is one of the most challenging tasks in surgical pathology because of its uncommon type (5-10% soft tissue sarcomas), morphologic resemblance to other spindle cell neoplasms and lack of sensitive and specific immunohistochemical markers. The pathologic diagnosis is more straightforward in the clinical setting of neurofibromatosis-1, but problems are mainly centered on the non-neurofibromatosis-1 malignant peripheral nerve sheath tumors. To date, S100 protein is the most widely applied marker in the case of a suspected malignant peripheral nerve sheath tumor, yet its suboptimal sensitivity and its expression in other spindle cell neoplasms, including spindle cell melanoma, clear-cell sarcoma, leiomyosarcoma and monophasic synovial sarcoma, add to the diagnostic conundrum. Growth-associated protein 43 (GAP43), a membrane-associated phosphoprotein expressed in neuronal growth cones and Schwann cell precursors during neural development and axonal regeneration, was applied to a set of nerve sheath and non-nerve sheath spindle cell neoplasms. The findings in this study indicate that GAP43 is expressed in malignant peripheral nerve sheath tumors (n=18/21; 86%) and demonstrates a sensitivity superior to S100 protein (n=13/21; 62%). GAP43 is also positive in neurofibromas (n=17/18; 94%), schwannomas (n=11/12; 92%) and desmoplastic melanomas (n=7/10; 70%). In contrast, it is negative in the non-desmoplastic spindle cell melanomas (n=20/22; 91%). Of the other non-neural soft tissue sarcomas, GAP43 is non-reactive in most leiomyosarcomas (n=14/16; 88%) and clear-cell sarcomas (n=8/8), and only focally positive in monophasic synovial sarcomas (n=3/7; 43%). GAP43 is seemingly a highly sensitive marker for peripheral nerve

  19. Angiogenic Effects of Dimeric Dipeptide Mimetic of Loop 4 of Nerve Growth Factor.

    PubMed

    Kryzhanovskii, S A; Antipova, T A; Tsorin, I B; Pekeldina, E S; Stolyaruk, V N; Nikolaev, S V; Sorokina, A V; Gudasheva, T A; Seredenin, S B

    2016-08-01

    Angiogenic action of compound GK-2, a dimeric dipeptide mimetic of loop 4 of nerve growth factor (NGF), was studied in in vitro and in vivo experiments. Experiments on human endothelial cell culture HUVEC showed that compound GK-2 significantly (p<0.05) stimulated the initial stage of angiogenesis, and its angiogenic activity was not inferior to the reference neurotrophin NGF. In experiments with hindlimb ischemia modeled in rats, GK-2 (1 mg/kg intraperitoneally for 14 days) significantly increased the total length of capillary vessels (p<0.003) and the number of vessels per 1 mm2 ischemic tissue (p<0.001) in comparison with the control. Our findings indicate that under experimental conditions compound GK-2 exhibits not only angiogenic, but also anti-ischemic activity.

  20. Antagonism of nerve growth factor-TrkA signaling and the relief of pain.

    PubMed

    Mantyh, Patrick W; Koltzenburg, Martin; Mendell, Lorne M; Tive, Leslie; Shelton, David L

    2011-07-01

    Nerve growth factor (NGF) was originally discovered as a neurotrophic factor essential for the survival of sensory and sympathetic neurons during development. However, in the adult NGF has been found to play an important role in nociceptor sensitization after tissue injury. The authors outline mechanisms by which NGF activation of its cognate receptor, tropomyosin-related kinase A receptor, regulates a host of ion channels, receptors, and signaling molecules to enhance acute and chronic pain. The authors also document that peripherally restricted antagonism of NGF-tropomyosin-related kinase A receptor signaling is effective for controlling human pain while appearing to maintain normal nociceptor function. Understanding whether there are any unexpected adverse events and how humans may change their behavior and use of the injured/degenerating tissue after significant pain relief without sedation will be required to fully appreciate the patient populations that may benefit from these therapies targeting NGF.

  1. Tiam1 as a signaling mediator of nerve growth factor-dependent neurite outgrowth.

    PubMed

    Shirazi Fard, Shahrzad; Kele, Julianna; Vilar, Marçal; Paratcha, Gustavo; Ledda, Fernanda

    2010-03-19

    Nerve Growth Factor (NGF)-induced neuronal differentiation requires the activation of members of the Rho family of small GTPases. However, the molecular mechanisms through which NGF regulates cytoskeletal changes and neurite outgrowth are not totally understood. In this work, we identify the Rac1-specific guanine exchange factor (GEF) Tiam1 as a novel mediator of NGF/TrkA-dependent neurite elongation. In particular, we report that knockdown of Tiam1 causes a significant reduction in Rac1 activity and neurite outgrowth induced by NGF. Physical interaction between Tiam1 and active Ras (Ras-GTP), but not tyrosine phosphorylation of Tiam1, plays a central role in Rac1 activation by NGF. In addition, our findings indicate that Ras is required to associate Tiam1 with Rac1 and promote Rac1 activation upon NGF stimulation. Taken together, these findings define a novel molecular mechanism through which Tiam1 mediates TrkA signaling and neurite outgrowth induced by NGF.

  2. 'Mature' nerve growth factor is a minor species in most peripheral tissues.

    PubMed

    Bierl, Michael A; Jones, Elizabeth E; Crutcher, Keith A; Isaacson, Lori G

    The classic neurotrophin hypothesis is based on the idea that innervating neurons derive 'mature' neurotrophin provided by the target for their survival. Yet large precursor forms of the neurotrophin nerve growth factor (NGF) have been reported in both central and peripheral tissues. In the present study, immunoblotting was used to survey peripheral tissues containing NGF-responsive neurons and to characterize various NGF species. These results demonstrate that 'mature' forms of NGF, i.e., the 13 and 16kDa species, are rare in sympathetic and sensory ganglia and in their peripheral targets, and that large molecular weight NGF precursors are abundant. In addition, certain NGF forms predominate in a given tissue, with each tissue exhibiting a characteristic NGF expression pattern. These findings suggest that NGF processing in peripheral tissues and in NGF-responsive ganglia may involve a variety of NGF species.

  3. Analytical modelling of retrograde transport of nerve growth factors in an axon: a transient problem.

    PubMed

    Kuznetsov, A V

    2013-01-01

    The purpose of this paper was to develop an analytical solution describing retrograde transport of nerve growth factors (NGFs) from a target tissue to the neuron soma. The obtained solution is applied to describe two situations: (1) when all dynein motors are moving at a constant velocity and (2) when the dynein velocity distribution is described by a probability density function. The dynamics of NGF concentrations and fluxes is investigated. It is established that the magnitude of the diffusion flux forms a wave localised in space and propagating towards the neuron soma; the magnitude of this wave decreases as the wave propagates downstream. The dynein-driven flux of NGFs is demonstrated to be the main component in the NGF flux, which is shown to be strongly correlated with the NGF concentration.

  4. Nerve growth factor is an autocrine factor essential for the survival of macrophages infected with HIV.

    PubMed

    Garaci, E; Caroleo, M C; Aloe, L; Aquaro, S; Piacentini, M; Costa, N; Amendola, A; Micera, A; Caliò, R; Perno, C F; Levi-Montalcini, R

    1999-11-23

    Nerve growth factor (NGF) is a neurotrophin with the ability to exert specific effects on cells of the immune system. Human monocytes/macrophages (M/M) infected in vitro with HIV type 1 (HIV-1) are able to produce substantial levels of NGF that are associated with enhanced expression of the high-affinity NGF receptor (p140 trkA) on the M/M surface. Treatment of HIV-infected human M/M with anti-NGF Ab blocking the biological activity of NGF leads to a marked decrease of the expression of p140 trkA high-affinity receptor, a concomitant increased expression of p75(NTR) low-affinity receptor for NGF, and the occurrence of apoptotic death of M/M. Taken together, these findings suggest a role for NGF as an autocrine survival factor that rescues human M/M from the cytopathic effect caused by HIV infection.

  5. Cortical peroxynitration of nerve growth factor in aged and cognitively impaired rats.

    PubMed

    Bruno, Martin A; Cuello, A Claudio

    2012-09-01

    Basal forebrain cholinergic neurons (BFCN), a system involved in learning and memory processes, are highly dependent on a continuous supply of biologically active nerve growth factor (NGF). Age-related cholinergic atrophy and cell loss in normal brains is apparently not complemented by reductions in the levels of NGF as could be expected. In the present work, cortical proNGF/NGF were immunoprecipitated from cortical brain homogenates from young and aged and behaviorally characterized rats and resolved with antinitrotyrosine antibodies to reveal nitration of tyrosine residues in proteins. Cortical proNGF in aged and cognitively impaired rats was found to be a target for peroxynitrite-mediated oxidative damage with correlative impact on decrease in choline acetyltransferase activity. These studies provide evidence for oxidative stress damage of NGF molecules in the cerebral cortex of cognitively impaired aged rats as previously shown in AD human brains. Copyright © 2012 Elsevier Inc. All rights reserved.

  6. Antagonism of Nerve Growth Factor-TrkA Signaling and the Relief of Pain

    PubMed Central

    Mantyh, Patrick W.; Koltzenburg, Martin; Mendell, Lorne M.; Tive, Leslie; Shelton, David L.

    2011-01-01

    Nerve growth factor (NGF) was originally discovered as a neurotrophic factor essential for the survival of sensory and sympathetic neurons during development. However in the adult, NGF has been found to play an important role in nociceptor sensitization following tissue injury. Here we outline mechanisms by which NGF activation of its cognate receptor, tropomyosin-related kinase A receptor, regulates a host of ion channels, receptors, and signaling molecules to enhance acute and chronic pain. Further, we document that peripherally restricted antagonism of NGF-tropomyosin-related kinase A receptor signaling is effective for controlling human pain while appearing to maintain normal nociceptor function. Understanding whether there are any unexpected adverse events as well as how humans may change their behavior and use of the injured/degenerating tissue following significant pain relief without sedation will be required to fully appreciate the patient populations that may benefit from these therapies targeting NGF. PMID:21602663

  7. Prophylactic pretreatment of mice with hematopoietic growth factors induces expansion of primitive cell compartments and results in protection against 5-fluorouracil-induced toxicity.

    PubMed

    de Haan, G; Donte, B; Engel, C; Loeffler, M; Nijhof, W

    1996-06-01

    The aim of this study was to expand the primitive and committed hematopoietic cell compartments in vivo in order to confer resistance of the blood cell forming system against the cytotoxic, cell cycle specific drug 5-fluorouracil (5-FU). Possible chemoprotective effects of such a pretreatment could result from increased numbers of hematopoietic cells, present before 5-FU administration. In addition, we hypothesized that an enhanced number of primitive and progenitor calls would result in a reduced cycling activity, ie, 5-FU sensitivity, of these same cells, due to normal physiological feedback loops. Administration of stem cell factor (SCF) plus interleukin-11 (IL-11) to mice was shown to result in expansion of the various immature cell compartments in marrow and, in particular, spleen. The total body content of the primitive cobblestone area forming cells (CAFC)-day 28 was increased to 140%, whereas the more committed cells (CAFC-day 7, erythroid and granuloid progenitors) were increased to 500%. This in vivo expansion resulted in a decreased 5-FU sensitivity of the hematopoietic system. In particular, mice that had received 5-FU 24 hours after discontinuation of growth factor pretreatment showed significantly less toxicity of committed cell stages. Compared with mice not pretreated, it appeared that in pretreated mice, 24 hours after 5-FU administration, the absolute number, but also the fraction of surviving CAFC, was much higher in both marrow and spleen. This was caused by a decrease in the cycling activity of all primitive cell subsets. To explore the possible use of this finding in a chemotherapeutic setting, we determined the interval between two subsequent doses of 5-FU (160 mg/kg) that was required to prevent drug-induced mortality. When control mice received a second dose of 5-FU 7, 10, or 14 days after the first, respectively 0%, 20%, and 80% survived. In contrast, 40% and 100% of mice that received SCF + IL-11 before the first dose of 5-FU, survived a

  8. Axonal regeneration and remyelination evaluation of chitosan/gelatin-based nerve guide combined with transforming growth factor-β1 and Schwann cells.

    PubMed

    Nie, Xin; Deng, Manjing; Yang, Maojin; Liu, Luchuan; Zhang, Yongjie; Wen, Xiujie

    2014-01-01

    Despite efforts in peripheral nerve injury and regeneration, it is difficult to achieve a functional recovery following extended peripheral nerve lesions. Even if artificial nerve conduit, cell components and growth factors can enhance nerve regeneration, integration in peripheral nerve repair and regeneration remains yet to be explored. For this study, we used chitosan/gelatin nerve graft constructed with collagenous matrices as a vehicle for Schwann cells and transforming growth factor-β1 to bridge a 10-mm gap of the sciatic nerve and explored the feasibility of improving regeneration and reinnervation in rats. The nerve regeneration was assessed with functional recovery, electrophysiological test, retrograde labeling, and immunohistochemistry analysis during the post-operative period of 16 weeks. The results showed that the internal sides of the conduits were compact enough to prevent the connective tissues from ingrowth. Nerve conduction velocity, average regenerated myelin area, and myelinated axon count were similar to those treated with autograft (p > 0.05) but significantly higher than those bridged with chitosan/gelatin nerve graft alone (p < 0.05). Evidences from retrograde labeling and immunohistochemistry analysis are further provided in support of improving axonal regeneration and remyelination. A designed graft incorporating all of the tissue-engineering strategies for peripheral nerve regeneration may provide great progress in tissue engineering for nerve repair.

  9. Nuclear translocation of p42/p44 mitogen-activated protein kinase is required for growth factor-induced gene expression and cell cycle entry.

    PubMed Central

    Brunet, A; Roux, D; Lenormand, P; Dowd, S; Keyse, S; Pouysségur, J

    1999-01-01

    Mitogen-activated protein kinase (MAPK) modules, composed of three protein kinases activated by successive phosphorylation, are involved in the signal transduction of a wide range of extracellular agents. In mammalian cells, mitogenic stimulation triggers the translocation of p42/p44MAPK from the cytoplasm to the nucleus, whereas the other protein kinases of the module remain cytosolic. Since MAPK has been shown to phosphorylate and activate nuclear targets, such as the transcription factor Elk1, it has been proposed, but not yet demonstrated, that MAPK nuclear translocation could represent a critical step in signal transduction. In this study, we sequestered p42/p44MAPK in the cytoplasm by the expression of a catalytically inactive form of cytoplasmic MAP kinase phosphatase (MKP-3/Pyst-1). Sequestering MAPK in the cytoplasm did not alter its activation or its ability to phosphorylate cytoplasmic substrates of MAPK (p90RSK1 or an engineered cytoplasmic form of Elk1). In contrast, prevention of MAPK nuclear translocation strongly inhibited Elk1-dependent gene transcription and the ability of cells to reinitiate DNA replication in response to growth factors. Thus the relocalization of MAPK to the nucleus appears to be an important regulatory step for mitogen-induced gene expression and cell cycle re-entry. PMID:9927426

  10. Basic fibroblast growth factor induces matrix metalloproteinase-13 via ERK MAP kinase-altered phosphorylation and sumoylation of Elk-1 in human adult articular chondrocytes.

    PubMed

    Im, Hee-Jeong; Sharrocks, Andrew D; Lin, Xia; Yan, Dongyao; Kim, Jaesung; van Wijnen, Andre J; Hipskind, Robert A

    2009-01-01

    Degradation of the extracellular matrix (ECM) by matrix metalloproteinases (MMPs) and release of basic fibroblast growth factor (bFGF) are principal aspects of the pathology of osteoarthritis (OA). ECM disruption leads to bFGF release, which activates the extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) pathway and its downstream target the Ets-like transcription factor Elk-1. Previously we demonstrated that the bFGF-ERK-Elk-1 signaling axis is responsible for the potent induction of MMP-13 in human primary articular chondrocytes. Here we report that, in addition to phosphorylation of Elk-1, dynamic posttranslational modification of Elk-1 by small ubiquitin-related modifier (SUMO) serves as an important mechanism through which MMP-13 gene expression is regulated. We show that bFGF activates Elk-1 mainly through the ERK pathway and that increased phosphorylation of Elk-1 is accompanied by decreased conjugation of SUMO to Elk-1. Reporter gene assays reveal that phosphorylation renders Elk-1 competent for induction of MMP-13 gene transcription, while sumoylation has the opposite effect. Furthermore, we demonstrate that the SUMO-conjugase Ubc9 acts as a key mediator for Elk-1 sumoylation. Taken together, our results suggest that sumoylation antagonizes the phosphorylation-dependent transactivation capacity of Elk-1. This attenuates transcription of its downstream target gene MMP-13 to maintain the integrity of cartilage ECM homeostasis.

  11. Eicosopentaneoic Acid and Other Free Fatty Acid Receptor Agonists Inhibit Lysophosphatidic Acid- and Epidermal Growth Factor-Induced Proliferation of Human Breast Cancer Cells.

    PubMed

    Hopkins, Mandi M; Zhang, Zhihong; Liu, Ze; Meier, Kathryn E

    2016-01-26

    Many key actions of ω-3 (n-3) fatty acids have recently been shown to be mediated by two G protein-coupled receptors (GPCRs) in the free fatty acid receptor (FFAR) family, FFA1 (GPR40) and FFA4 (GPR120). n-3 Fatty acids inhibit proliferation of human breast cancer cells in culture and in animals. In the current study, the roles of FFA1 and FFA4 were investigated. In addition, the role of cross-talk between GPCRs activated by lysophosphatidic acid (LPA), and the tyrosine kinase receptor activated by epidermal growth factor (EGF), was examined. In MCF-7 and MDA-MB-231 human breast cancer cell lines, both LPA and EGF stimulated proliferation, Erk activation, Akt activation, and CCN1 induction. LPA antagonists blocked effects of LPA and EGF on proliferation in MCF-7 and MDA-MB-231, and on cell migration in MCF-7. The n-3 fatty acid eicosopentaneoic acid inhibited LPA- and EGF-induced proliferation in both cell lines. Two synthetic FFAR agonists, GW9508 and TUG-891, likewise inhibited LPA- and EGF-induced proliferation. The data suggest a major role for FFA1, which was expressed by both cell lines. The results indicate that n-3 fatty acids inhibit breast cancer cell proliferation via FFARs, and suggest a mechanism involving negative cross-talk between FFARS, LPA receptors, and EGF receptor.

  12. Eicosopentaneoic Acid and Other Free Fatty Acid Receptor Agonists Inhibit Lysophosphatidic Acid- and Epidermal Growth Factor-Induced Proliferation of Human Breast Cancer Cells

    PubMed Central

    Hopkins, Mandi M.; Zhang, Zhihong; Liu, Ze; Meier, Kathryn E.

    2016-01-01

    Many key actions of ω-3 (n-3) fatty acids have recently been shown to be mediated by two G protein-coupled receptors (GPCRs) in the free fatty acid receptor (FFAR) family, FFA1 (GPR40) and FFA4 (GPR120). n-3 Fatty acids inhibit proliferation of human breast cancer cells in culture and in animals. In the current study, the roles of FFA1 and FFA4 were investigated. In addition, the role of cross-talk between GPCRs activated by lysophosphatidic acid (LPA), and the tyrosine kinase receptor activated by epidermal growth factor (EGF), was examined. In MCF-7 and MDA-MB-231 human breast cancer cell lines, both LPA and EGF stimulated proliferation, Erk activation, Akt activation, and CCN1 induction. LPA antagonists blocked effects of LPA and EGF on proliferation in MCF-7 and MDA-MB-231, and on cell migration in MCF-7. The n-3 fatty acid eicosopentaneoic acid inhibited LPA- and EGF-induced proliferation in both cell lines. Two synthetic FFAR agonists, GW9508 and TUG-891, likewise inhibited LPA- and EGF-induced proliferation. The data suggest a major role for FFA1, which was expressed by both cell lines. The results indicate that n-3 fatty acids inhibit breast cancer cell proliferation via FFARs, and suggest a mechanism involving negative cross-talk between FFARS, LPA receptors, and EGF receptor. PMID:26821052

  13. UAP56 is an important mediator of Angiotensin II/platelet derived growth factor induced vascular smooth muscle cell DNA synthesis and proliferation

    SciTech Connect

    Sahni, Abha; Wang, Nadan; Alexis, Jeffrey

    2013-02-15

    Highlights: ► Knockdown of UAP56 inhibits Angiotensin II/PDGF induced vascular smooth muscle cell proliferation. ► UAP56 is a positive regulator of E2F transcriptional activation. ► UAP56 is present in the vessel wall of low flow carotid arteries. -- Abstract: Angiotensin (Ang) II and platelet-derived growth factor (PDGF) are important mediators of pathologic vascular smooth muscle cell (VSMC) proliferation. Identifying downstream mediators of Ang II and PDGF signaling may provide insights for therapies to improve vascular proliferative diseases. We have previously demonstrated that breakpoint cluster region (Bcr) is an important mediator of Ang II/PDGF signaling in VSMC. We have recently reported that the DExD/H box protein UAP56 is an interacting partner of Bcr in regulating VSMC DNA synthesis. We hypothesized that UAP56 itself is an important regulator of VSMC proliferation. In this report we demonstrate that knockdown of UAP56 inhibits Ang II/PDGF induced VSMC DNA synthesis and proliferation, and inhibits E2F transcriptional activity. In addition, we demonstrate that UAP56 is present in the vessel wall of low-flow carotid arteries. These findings suggest that UAP56 is a regulator of VSMC proliferation and identify UAP56 as a target for preventing vascular proliferative disease.

  14. Dihydroartemisinin inhibits vascular endothelial growth factor-induced endothelial cell migration by a p38 mitogen-activated protein kinase-independent pathway.

    PubMed

    Guo, Ling; Dong, Fengyun; Hou, Yinglong; Cai, Weidong; Zhou, Xia; Huang, Ai-Ling; Yang, Min; Allen, Thaddeus D; Liu, Ju

    2014-12-01

    Dihydroartemisinin (DHA), a semi-synthetic derivative of artemisinin, has been demonstrated to possess a strong antiangiogenic activity. However, the molecular mechanisms underlying this effect remain unclear. Endothelial cell (EC) migration is an essential component of angiogenesis, and the p38 mitogen-activated protein kinase (MAPK) signaling pathway plays a key role in the regulation of migration induced by vascular endothelial growth factor (VEGF). The aim of the present study was to investigate the effects of DHA on EC migration and the p38 MAPK signaling pathway. Human umbilical vein ECs (HUVECs) were treated with DHA and VEGF-induced migration was analyzed. The activation of p38 MAPK was detected by western blot analysis, and the migration assays were performed with a p38-specific inhibitor, SB203850. It was revealed that 20 μM DHA significantly reduced EC migration in the transwell migration assay, wound healing assay and electrical cell-substrate impedance sensing real-time analysis. However, DHA did not affect p38 MAPK phosphorylation or expression. In the absence or presence of SB203850, DHA induced a similar proportional reduction of EC migration in the three migration assays. Therefore, the present study demonstrated that DHA inhibits VEGF-induced EC migration via a p38 MAPK-independent pathway.

  15. Dual regulation of cofilin activity by LIM kinase and Slingshot-1L phosphatase controls platelet-derived growth factor-induced migration of human aortic smooth muscle cells.

    PubMed

    San Martín, Alejandra; Lee, Moo Yeol; Williams, Holly C; Mizuno, Kensaku; Lassègue, Bernard; Griendling, Kathy K

    2008-02-29

    Platelet-derived growth factor (PDGF) plays a central role in vascular healing, atherosclerosis, and restenosis, partly by stimulating vascular smooth muscle cell (VSMC) migration. Migration requires rapid turnover of actin filaments, which is partially controlled by cofilin. Although cofilin is negatively regulated by Ser3 phosphorylation, the upstream signaling pathways have not been defined, nor has its role in VSMC migration been studied. We hypothesized that PDGF-induced migration of VSMCs involves cofilin activation and that this is regulated by the serine kinase LIM kinase (LIMK) and the novel phosphatase Slingshot (SSH)1L. In human VSMCs, stimulation with PDGF increased G-actin incorporation into the actin cytoskeleton. PDGF transiently activated the cofilin kinase, LIMK, with a peak at 5 minutes. However, cofilin was dephosphorylated between 5 and 45 minutes, with a maximum of 43+/-5% dephosphorylation at 30 minutes, suggesting that PDGF also activates a cofilin phosphatase. We found that VSMCs express SSH1L, which is induced and activated (564+/-73 versus 1021+/-141 picomoles of PO(4); P=0.015) by PDGF. Of importance, small interfering RNA directed against SSH1L blocked cofilin dephosphorylation and decreased migration (528+/-33 versus 318+/-25 cells/field; P<0.01). Taken together, our results suggest that PDGF participates in actin dynamics by dual regulation of cofilin activity via LIMK and SSH1L.

  16. Gelsolin-mediated activation of PI3K/Akt pathway is crucial for hepatocyte growth factor-induced cell scattering in gastric carcinoma

    PubMed Central

    Huang, Baohua; Deng, Shuo; Loo, Ser Yue; Datta, Arpita; Yap, Yan Lin; Yan, Benedict; Ooi, Chia Huey; Dinh, Thuy Duong; Zhuo, Jingli; Tochhawng, Lalchhandami; Gopinadhan, Suma; Jegadeesan, Tamilarasi; Tan, Patrick; Salto-Tellez, Manuel; Yong, Wei Peng; Soong, Richie; Yeoh, Khay Guan; Goh, Yaw Chong; Lobie, Peter E.; Yang, Henry; Kumar, Alan Prem; Maciver, Sutherland K.; So, Jimmy B.Y.; Yap, Celestial T.

    2016-01-01

    In gastric cancer (GC), the main subtypes (diffuse and intestinal types) differ in pathological characteristics, with diffuse GC exhibiting early disseminative and invasive behaviour. A distinctive feature of diffuse GC is loss of intercellular adhesion. Although widely attributed to mutations in the CDH1 gene encoding E-cadherin, a significant percentage of diffuse GC do not harbor CDH1 mutations. We found that the expression of the actin-modulating cytoskeletal protein, gelsolin, is significantly higher in diffuse-type compared to intestinal-type GCs, using immunohistochemical and microarray analysis. Furthermore, in GCs with wild-type CDH1, gelsolin expression correlated inversely with CDH1 gene expression. Downregulating gelsolin using siRNA in GC cells enhanced intercellular adhesion and E-cadherin expression, and reduced invasive capacity. Interestingly, hepatocyte growth factor (HGF) induced increased gelsolin expression, and gelsolin was essential for HGF-medicated cell scattering and E-cadherin transcriptional repression through Snail, Twist and Zeb2. The HGF-dependent effect on E-cadherin was found to be mediated by interactions between gelsolin and PI3K-Akt signaling. This study reveals for the first time a function of gelsolin in the HGF/cMet oncogenic pathway, which leads to E-cadherin repression and cell scattering in gastric cancer. Our study highlights gelsolin as an important pro-disseminative factor contributing to the aggressive phenotype of diffuse GC. PMID:27058427

  17. Hepatocyte growth factor induces glucose uptake in 3T3-L1 adipocytes through A Gab1/phosphatidylinositol 3-kinase/Glut4 pathway.

    PubMed

    Bertola, Adeline; Bonnafous, Stéphanie; Cormont, Mireille; Anty, Rodolphe; Tanti, Jean-François; Tran, Albert; Le Marchand-Brustel, Yannick; Gual, Philippe

    2007-04-06

    Adipose tissue is a source of hepatocyte growth factor (HGF), and circulating HGF levels have been associated with elevated body mass index in human. However, the effects of HGF on adipocyte functions have not yet been investigated. We show here that in 3T3-L1 adipocytes HGF stimulates the phosphatidylinositol (PI) 3-kinase-dependent protein kinase B (PKB) activity, AS160 phosphorylation, Glut4 translocation, and consequently, glucose uptake. The initial steps involved in HGF- and insulin-induced glucose uptake are different. HGF enhanced the tyrosine phosphorylation of Gab1, leading to the recruitment of the p85-regulated subunit of PI 3-kinase, whereas p85 was exclusively recruited by IRS1 in response to insulin. In adipocytes rendered insulin-resistant by a long-lasting tumor necrosis factor alpha treatment, the protein level of Gab1 was strongly decreased, and HGF-stimulated PKB activation and glucose uptake were also altered. Moreover, treatment of 3T3-L1 adipocytes with thiazolidinedione, an anti-diabetic drug, enhanced the expression of both HGF and its receptor. These data provide the first evidence that in vitro HGF promotes glucose uptake through a Gab1/PI 3-kinase/PKB/AS160 pathway which was altered in tumor necrosis factor alpha-treated adipocytes.

  18. The epidermal growth factor-induced migration of rat liver epithelial cells is associated with a transient inhibition of DNA synthesis.

    PubMed

    Geimer, P; Bade, E G

    1991-10-01

    Epidermal growth factor (EGF) is a potent mitogen for most cultured cells and has previously been shown to induce the migration of rat liver epithelial cells. We have now demonstrated that under migration-inducing conditions EGF does not stimulate cell proliferation, but causes instead a transient inhibition of DNA synthesis. Analysis at the single-cell level by [3H]thymidine autoradiography indicated that in 40-50% of the EGF-treated cell population the entry into S phase is delayed. The simultaneous demonstration of migration tracks by laminin immunofluorescence revealed that the transient inhibition of DNA synthesis is not restricted to the migratory cells. The effect is also observed with the stationary subpopulation and appears, therefore, to be independent of the induction of migration. The independence of both processes was further supported by showing that induction of migration by EGF proceeds undisturbed in cells blocked in S phase by aphidicolin. These results indicated that for rat liver epithelial cells the induction of migration by EGF has priority over cell proliferation. The data also emphasize the need for a time-course analysis when studying factors that stimulate or inhibit DNA synthesis or cell proliferation.

  19. Involvement of cysteine-rich protein 61 in the epidermal growth factor-induced migration of human anaplastic thyroid cancer cells.

    PubMed

    Chin, Li-Han; Hsu, Sung-Po; Zhong, Wen-Bin; Liang, Yu-Chih

    2016-05-01

    Anaplastic thyroid cancer (ATC) is among the most aggressive types of malignant cancer. Epidermal growth factor (EGF) plays a crucial role in the pathogenesis of ATC, and patients with thyroid carcinoma typically exhibit increased cysteine-rich protein 61 (Cyr61). In this study, we found that EGF treatment induced cell migration, stress fiber formation, Cyr61 mRNA and protein expressions, and Cyr61 protein secretion in ATC cells. The recombinant Cyr61 protein significantly induced cell migration; however, inhibition of Cyr61 activity by a Cyr61-specific antibody abrogated EGF-induced cell migration. EGF treatment also affected epithelial-to-mesenchymal transition (EMT)-related marker protein expression, as evidenced by an increase in vimentin and a decrease in E-cadherin expression. Inhibition of Cyr61 expression by Cyr61 siRNA decreased cell migration and reversed the EMT-related marker protein expression. EGF treatment increased the phosphorylation of the extracellular signal-regulated kinase (ERK) and cAMP response element-binding protein (CREB), and finally activated Cyr61 promoter plasmid activity. Our results suggest that Cyr61 is induced by EGF through the ERK/CREB signal pathway and that it plays a crucial role in the migration and invasion of ATC cells; moreover, Cyr61 might be a therapeutic target for metastatic ATC.

  20. miR-503 inhibits platelet-derived growth factor-induced human aortic vascular smooth muscle cell proliferation and migration through targeting the insulin receptor.

    PubMed

    Bi, Rui; Ding, Fangbao; He, Yi; Jiang, Lianyong; Jiang, Zhaolei; Mei, Ju; Liu, Hao

    2016-12-01

    Abnormal proliferation and migration of vascular smooth muscle cells (VSMC) is a common feature of disease progression in atherosclerosis. Here, we investigated the potential role of miR-503 in platelet-derived growth factor (PDGF)-induced proliferation and migration of human aortic smooth muscle cells and the underlying mechanisms of action. miR-503 expression was significantly downregulated in a dose- and time-dependent manner following PDGF treatment. Introduction of miR-503 mimics into cultured SMCs significantly attenuated cell proliferation and migration induced by PDGF. Bioinformatics analyses revealed that the insulin receptor (INSR) is a target candidate of miR-503. miR-503 suppressed luciferase activity driven by a vector containing the 3'-untranslated region of INSR in a sequence-specific manner. Downregulation of INSR appeared critical for miR-503-mediated inhibitory effects on PDGF-induced cell proliferation and migration in human aortic SMCs. Based on the collective data, we suggest a novel role of miR-503 as a regulator of VSMC proliferation and migration through modulating INSR.

  1. Gelsolin-mediated activation of PI3K/Akt pathway is crucial for hepatocyte growth factor-induced cell scattering in gastric carcinoma.

    PubMed

    Huang, Baohua; Deng, Shuo; Loo, Ser Yue; Datta, Arpita; Yap, Yan Lin; Yan, Benedict; Ooi, Chia Huey; Dinh, Thuy Duong; Zhuo, Jingli; Tochhawng, Lalchhandami; Gopinadhan, Suma; Jegadeesan, Tamilarasi; Tan, Patrick; Salto-Tellez, Manuel; Yong, Wei Peng; Soong, Richie; Yeoh, Khay Guan; Goh, Yaw Chong; Lobie, Peter E; Yang, Henry; Kumar, Alan Prem; Maciver, Sutherland K; So, Jimmy B Y; Yap, Celestial T

    2016-05-03

    In gastric cancer (GC), the main subtypes (diffuse and intestinal types) differ in pathological characteristics, with diffuse GC exhibiting early disseminative and invasive behaviour. A distinctive feature of diffuse GC is loss of intercellular adhesion. Although widely attributed to mutations in the CDH1 gene encoding E-cadherin, a significant percentage of diffuse GC do not harbor CDH1 mutations. We found that the expression of the actin-modulating cytoskeletal protein, gelsolin, is significantly higher in diffuse-type compared to intestinal-type GCs, using immunohistochemical and microarray analysis. Furthermore, in GCs with wild-type CDH1, gelsolin expression correlated inversely with CDH1 gene expression. Downregulating gelsolin using siRNA in GC cells enhanced intercellular adhesion and E-cadherin expression, and reduced invasive capacity. Interestingly, hepatocyte growth factor (HGF) induced increased gelsolin expression, and gelsolin was essential for HGF-medicated cell scattering and E-cadherin transcriptional repression through Snail, Twist and Zeb2. The HGF-dependent effect on E-cadherin was found to be mediated by interactions between gelsolin and PI3K-Akt signaling. This study reveals for the first time a function of gelsolin in the HGF/cMet oncogenic pathway, which leads to E-cadherin repression and cell scattering in gastric cancer. Our study highlights gelsolin as an important pro-disseminative factor contributing to the aggressive phenotype of diffuse GC.

  2. Essential role for vav Guanine nucleotide exchange factors in brain-derived neurotrophic factor-induced dendritic spine growth and synapse plasticity.

    PubMed

    Hale, Carly F; Dietz, Karen C; Varela, Juan A; Wood, Cody B; Zirlin, Benjamin C; Leverich, Leah S; Greene, Robert W; Cowan, Christopher W

    2011-08-31

    Brain-derived neurotrophic factor (BDNF) and its cognate receptor, TrkB, regulate a wide range of cellular processes, including dendritic spine formation and functional synapse plasticity. However, the signaling mechanisms that link BDNF-activated TrkB to F-actin remodeling enzymes and dendritic spine morphological plasticity remain poorly understood. We report here that BDNF/TrkB signaling in neurons activates the Vav family of Rac/RhoA guanine nucleotide exchange factors through a novel TrkB-dependent mechanism. We find that Vav is required for BDNF-stimulated Rac-GTP production in cortical and hippocampal neurons. Vav is partially enriched at excitatory synapses in the postnatal hippocampus but does not appear to be required for normal dendritic spine density. Rather, we observe significant reductions in both BDNF-induced, rapid, dendritic spine head growth and in CA3-CA1 theta burst-stimulated long-term potentiation in Vav-deficient mouse hippocampal slices, suggesting that Vav-dependent regulation of dendritic spine morphological plasticity facilitates normal functional synapse plasticity.

  3. Transcriptional mechanism of vascular endothelial growth factor-induced expression of protein kinase CβII in chronic lymphocytic leukaemia cells

    PubMed Central

    Al-Sanabra, Ola; Duckworth, Andrew D.; Glenn, Mark A.; Brown, Benjamin R. B.; Angelillo, Piera; Lee, Kelvin; Herbert, John; Falciani, Francesco; Kalakonda, Nagesh; Slupsky, Joseph R.

    2017-01-01

    A key feature of chronic lymphocytic leukaemia (CLL) cells is overexpressed protein kinase CβII (PKCβII), an S/T kinase important in the pathogenesis of this and other B cell malignancies. The mechanisms contributing to enhanced transcription of the gene coding for PKCβII, PRKCB, in CLL cells remain poorly described, but could be important because of potential insight into how the phenotype of these cells is regulated. Here, we show that SP1 is the major driver of PKCβII expression in CLL cells where enhanced association of this transcription factor with the PRKCB promoter is likely because of the presence of histone marks permissive of gene activation. We also show how vascular endothelial growth factor (VEGF) regulates PRKCB promoter function in CLL cells, stimulating PKCβ gene transcription via increased association of SP1 and decreased association of STAT3. Taken together, these results are the first to demonstrate a clear role for SP1 in the up regulation of PKCβII expression in CLL cells, and the first to link SP1 with the pathogenesis of this and potentially other B cell malignancies where PKCβII is overexpressed. PMID:28233872

  4. Filamin B Plays a Key Role in Vascular Endothelial Growth Factor-induced Endothelial Cell Motility through Its Interaction with Rac-1 and Vav-2*

    PubMed Central

    del Valle-Pérez, Beatriz; Martínez, Vanesa Gabriela; Lacasa-Salavert, Cristina; Figueras, Agnès; Shapiro, Sandor S.; Takafuta, Toshiro; Casanovas, Oriol; Capellà, Gabriel; Ventura, Francesc; Viñals, Francesc

    2010-01-01

    Actin-binding proteins filamin A (FLNA) and B (FLNB) are expressed in endothelial cells and play an essential role during vascular development. In order to investigate their role in adult endothelial cell function, we initially confirmed their expression pattern in different adult mouse tissues and cultured cell lines and found that FLNB expression is concentrated mainly in endothelial cells, whereas FLNA is more ubiquitously expressed. Functionally, small interfering RNA knockdown of endogenous FLNB in human umbilical vein endothelial cells inhibited vascular endothelial growth factor (VEGF)-induced in vitro angiogenesis by decreasing endothelial cell migration capacity, whereas FLNA ablation did not alter these parameters. Moreover, FLNB-depleted cells increased their substrate adhesion with more focal adhesions. The molecular mechanism underlying this effect implicates modulation of small GTP-binding protein Rac-1 localization and activity, with altered activation of its downstream effectors p21 protein Cdc42/Rac-activated kinase (PAK)-4/5/6 and its activating guanine nucleotide exchange factor Vav-2. Moreover, our results suggest the existence of a signaling complex, including FLNB, Rac-1, and Vav-2, under basal conditions that would further interact with VEGFR2 and integrin αvβ5 after VEGF stimulation. In conclusion, our results reveal a crucial role for FLNB in endothelial cell migration and in the angiogenic process in adult endothelial cells. PMID:20110358

  5. Nerve growth factor partially recovers inflamed skin from stress-induced worsening in allergic inflammation.

    PubMed

    Peters, Eva M J; Liezmann, Christiane; Spatz, Katharina; Daniltchenko, Maria; Joachim, Ricarda; Gimenez-Rivera, Andrey; Hendrix, Sven; Botchkarev, Vladimir A; Brandner, Johanna M; Klapp, Burghard F

    2011-03-01

    Neuroimmune dysregulation characterizes atopic disease, but its nature and clinical impact remain ill-defined. Induced by stress, the neurotrophin nerve growth factor (NGF) may worsen cutaneous inflammation. We therefore studied the role of NGF in the cutaneous stress response in a mouse model for atopic dermatitis-like allergic dermatitis (AlD). Combining several methods, we found that stress increased cutaneous but not serum or hypothalamic NGF in telogen mice. Microarray analysis showed increased mRNAs of inflammatory and growth factors associated with NGF in the skin. In stress-worsened AlD, NGF-neutralizing antibodies markedly reduced epidermal thickening together with NGF, neurotrophin receptor (tyrosine kinase A and p75 neurotrophin receptor), and transforming growth factor-β expression by keratinocytes but did not alter transepidermal water loss. Moreover, NGF expression by mast cells was reduced; this corresponded to reduced cutaneous tumor necrosis factor-α (TNF-α) mRNA levels but not to changes in mast cell degranulation or in the T helper type 1 (Th1)/Th2 cytokine balance. Also, eosinophils expressed TNF receptor type 2, and we observed reduced eosinophil infiltration after treatment with NGF-neutralizing antibodies. We thus conclude that NGF acts as a local stress mediator in perceived stress and allergy and that increased NGF message contributes to worsening of cutaneous inflammation mainly by enhancing epidermal hyperplasia, pro-allergic cytokine induction, and allergy-characteristic cellular infiltration.

  6. The vascular endothelial growth factor-induced disruption of gap junctions is relayed by an autocrine communication via ATP release in coronary capillary endothelium.

    PubMed

    Thuringer, Dominique

    2004-12-01

    Little is known concerning how the coordination of Ca(2+) signaling aids in capillary endothelial cell (CEC) functions, such as microvascular permeability and angiogenesis. Previous reports support the major involvement of gap junction (GJ) channels. However, the cell-to-cell communication may not be straightforward, especially if we consider the participation of active molecules released by CEC. In this study, short-term effects of vascular endothelial growth factor (VEGF-165) were compared with those of bradykinin (BK) on gap junction coupling (GJC) and remodeling of connexin-43 (Cx43) and then analyzed for intercellular Ca(2+) signal in primary cultures of coronary CEC. Dye-coupling experiments revealed that BK or VEGF completely blocked GJC. These effects correlated with the rapid internalization of Cx43 and its tyrosine phosphorylation in part via the phosphatidylinositol 3-kinase/Akt pathway. GJC slowly recovered with BK but not with VEGF in the following hour. In control conditions, mechanical stimulation of a single cell within a confluent monolayer triggered an intercellular Ca(2+) wave that was partially inhibited by GJC blockers or purinergic inhibitors. No wave propagation was observed after blockage of both GJC and purinergic receptors. Cell treatment with VEGF also reduced propagation of the Ca(2+) wave, which was totally prevented by applying a purinergic receptor antagonist but not with a GJC blocker. That excludes purine efflux through Cx hemichannels. We conclude that VEGF-induced disruption of GJC via Cx43 remodeling is relayed by an autocrine communication via secretion of ATP to preserve intercellular Ca(2+) signaling in capillary endothelium.

  7. The role of peroxisome proliferator-activated receptor-{beta}/{delta} in epidermal growth factor-induced HaCaT cell proliferation

    SciTech Connect

    Liang Pengfei; Jiang Bimei; Yang Xinghua; Xiao Xianzhong Huang Xu; Long Jianhong; Zhang Pihong; Zhang Minghua; Xiao Muzhang; Xie Tinghong; Huang Xiaoyuan

    2008-10-15

    Epidermal growth factor (EGF) has been shown to be a potent mitogen for epidermal cells both in vitro and in vivo, thus contributing to the development of an organism. It has recently become clear that peroxisome proliferator-activated receptor-{beta}/{delta} (PPAR{beta}/{delta}) expression and activation is involved in the cell proliferation. However, little is known about the role of PPAR{beta}/{delta} in EGF-induced proliferation of HaCaT keratinocytes. In this study, HaCaT cells were cultured in the presence and absence of EGF and we identified that EGF induced an increase of PPAR{beta}/{delta} mRNA and protein level expression in time-dependent and dose-dependent manner, and AG1487, an EGF receptor (EGFR) special inhibitor, caused attenuation of PPAR{beta}/{delta} protein expression. Electrophoretic mobility shift assay (EMSA) revealed that EGF significantly increased PPAR{beta}/{delta} binding activity in HaCaT keratinocytes. Antisense phosphorothioate oligonucleotides (asODNs) against PPAR{beta}/{delta} caused selectively inhibition of PPAR{beta}/{delta} protein content induced by EGF and significantly attenuated EGF-mediated cell proliferation. Treatment of the cells with L165041, a specific synthetic ligand for PPAR{beta}/{delta}, significantly enhanced EGF-mediated cell proliferation. Finally, c-Jun ablation inhibited PPAR{beta}/{delta} up-regulation induced by EGF, and chromatin immunoprecipitation (ChIP) showed that c-Jun bound to the PPAR{beta}/{delta} promoter and the binding increased in EGF-stimulated cells. These results demonstrate that EGF induces PPAR{beta}/{delta} expression in a c-Jun-dependent manner and PPAR{beta}/{delta} plays a vital role in EGF-stimulated proliferation of HaCaT cells.

  8. The inhibitory effect of dexamethasone on platelet-derived growth factor-induced vascular smooth muscle cell migration through up-regulating PGC-1{alpha} expression

    SciTech Connect

    Xu, Wei; Guo, Ting; Zhang, Yan; Jiang, Xiaohong; Zhang, Yongxian; Zen, Ke; Yu, Bo; Zhang, Chen-Yu

    2011-05-01

    Dexamethasone has been shown to inhibit vascular smooth muscle cell (VSMC) migration, which is required for preventing restenosis. However, the mechanism underlying effect of dexamethasone remains unknown. We have previously demonstrated that peroxisome proliferator-activated receptor gamma (PPAR{gamma}) coactivator-1 alpha (PGC-1{alpha}) can inhibit VSMC migration and proliferation. Here, we investigated the role of PGC-1{alpha} in dexamethasone-reduced VSMC migration and explored the possible mechanism. We first examined PGC-1{alpha} expression in cultured rat aortic VSMCs. The results revealed that incubation of VSMCs with dexamethasone could significantly elevate PGC-1{alpha} mRNA expression. In contrast, platelet-derived growth factor (PDGF) decreased PGC-1{alpha} expression while stimulating VSMC migration. Mechanistic study showed that suppression of PGC-1{alpha} by small interfering RNA strongly abrogated the inhibitory effect of dexamethasone on VSMC migration, whereas overexpression of PGC-1{alpha} had the opposite effect. Furthermore, an analysis of MAPK signal pathways showed that dexamethasone inhibited ERK and p38 MAPK phosphorylation in VSMCs. Overexpression of PGC-1{alpha} decreased both basal and PDGF-induced p38 MAPK phosphorylation, but it had no effect on ERK phosphorylation. Finally, inhibition of PPAR{gamma} activation by a PPAR{gamma} antagonist GW9662 abolished the suppressive effects of PGC-1{alpha} on p38 MAPK phosphorylation and VSMC migration. These effects of PGC-1{alpha} were enhanced by a PPAR{gamma} agonist troglitazone. Collectively, our data indicated for the first time that one of the anti-migrated mechanisms of dexamethasone is due to the induction of PGC-1{alpha} expression. PGC-1{alpha} suppresses PDGF-induced VSMC migration through PPAR{gamma} coactivation and, consequently, p38 MAPK inhibition.

  9. Egr-1 mediates epidermal growth factor-induced downregulation of E-cadherin expression via Slug in human ovarian cancer cells.

    PubMed

    Cheng, J-C; Chang, H-M; Leung, P C K

    2013-02-21

    Loss of the cell adhesion protein E-cadherin increases the invasive capability of ovarian cancer cells. We have previously shown that epidermal growth factor (EGF) downregulates E-cadherin and induces ovarian cancer cell invasion through the H(2)O(2)/p38 MAPK-mediated upregulation of the E-cadherin transcriptional repressor Snail. However, the molecular mechanisms underlying the EGF-induced downregulation of E-cadherin are not fully understood. In the current study, we demonstrated that treatment of two ovarian cancer cell lines, SKOV3 and OVCAR5, with EGF induced the expression of the transcription factor Egr-1, and this induction was abolished by small interfering RNA (siRNA)-mediated depletion of the EGF receptor. EGF-induced Egr-1 expression required the activation of the ERK1/2 and PI3K/Akt signaling pathways and was unrelated to EGF-induced H(2)O(2) production and activation of the p38 MAPK pathway. Moreover, depletion of Egr-1 with siRNA abolished the EGF-induced downregulation of E-cadherin and increased cell invasion. Interestingly, siRNA depletion of Egr-1 attenuated the EGF-induced expression of Slug, but not that of Snail. Moreover, chromatin immunoprecipitation (ChIP) analysis showed that Slug is a target gene of Egr-1. These results provide evidence that Egr-1 is a mediator that is involved in the EGF-induced downregulation of E-cadherin and increased cell invasion. Our results also demonstrate that EGF activates two independent signaling pathways, which are the H(2)O(2)/p38 MAPK-mediated upregulation of Snail expression and the Egr-1-mediated upregulation of Slug expression. These two signaling pathways contribute to the EGF-induced downregulation of E-cadherin, which subsequently increases the invasive capability of ovarian cancer cells.

  10. Hypoxia-inducible factor 1 alpha mediates epidermal growth factor-induced down-regulation of E-cadherin expression and cell invasion in human ovarian cancer cells.

    PubMed

    Cheng, Jung-Chien; Klausen, Christian; Leung, Peter C K

    2013-02-28

    Hypoxia-inducible factor 1α (HIF-1α) regulates the transcription of a number of genes under hypoxia and other extracellular stimulations. It has been shown that E-cadherin is down-regulated by epidermal growth factor receptor (EGF) stimulation, and that cells with low E-cadherin expression are more invasive. Our recent study demonstrated a novel mechanism by which EGF down-regulates E-cadherin expression through production of hydrogen peroxide (H(2)O(2)) and the activation of p38 MAPK in human ovarian cancer cells. In this study, we were interested in examining the potential role of HIF-1α in cell invasion under normoxic conditions, specifically when cells are treated with EGF, which is known to down-regulate E-cadherin and increase invasiveness. We show that EGF treatment induces HIF-1α expression in two human ovarian cancer cell lines (SKOV3 and OVCAR5), and that this effect is diminished by treatment with a membrane-permeable H(2)O(2) scavenger, PEG-catalase. However, the induction of HIF-1α by EGF did not require the activation of p38 MAPK. Treatment with siRNA targeting HIF-1α reduces both basal and EGF-induced HIF-1α levels. Importantly, treatment with HIF-1α siRNA diminishes the up-regulation of Snail and Slug as well as the down-regulation of E-cadherin by EGF. The involvement of HIF-1α in the down-regulation of E-cadherin was confirmed with cobalt chloride (CoCl(2)), a hypoxia-mimetic reagent. Finally, we also show that EGF-induced cell invasion is attenuated by treatment with HIF-1α siRNA. This study demonstrates an important role for HIF-1α in mediating the effects of EGF on Snail, Slug and E-cadherin expression as well as invasiveness in human ovarian cancer cells.

  11. 5-Lipoxygenase and cysteinyl leukotriene receptor 1 regulate epidermal growth factor-induced cell migration through Tiam1 upregulation and Rac1 activation.

    PubMed

    Magi, Shigeyuki; Takemoto, Yasushi; Kobayashi, Hiroki; Kasamatsu, Masato; Akita, Takahiro; Tanaka, Ayako; Takano, Kei; Tashiro, Etsu; Igarashi, Yasuhiro; Imoto, Masaya

    2014-03-01

    Cell migration is an essential step for tumor metastasis. The small GTPase Rac1 plays an important role in cell migration. Previously, we reported that epidermal growth factor (EGF) induced two waves of Rac1 activation; namely, at 5 min and 12 h after stimulation. A second wave of EGF-induced Rac1 activation was required for EGF-induced cell migration, however, the spatiotemporal regulation of the second wave of EGF-induced Rac1 activation remains largely unclear. In this study, we found that 5-lipoxygenase (5-LOX) is activated in the process of EGF-induced cell migration, and that leukotriene C4 (LTC4 ) produced by 5-LOX mediated the second wave of Rac1 activation, as well as cell migration. Furthermore, these effects caused by LTC4 were found to be blocked in the presence of the antagonist of cysteinyl leukotriene receptor 1 (CysLT1). This blockage indicates that LTC4 -mediated CysLT1 signaling regulates the second EGF-induced wave of Rac1 activation. We also found that 5-LOX inhibitors, CysLT1 antagonists and the knockdown of CysLT1 inhibited EGF-induced T cell lymphoma invasion and metastasis-inducing protein 1 (Tiam1) expression. Tiam1 expression is required for the second wave of EGF-induced Rac1 activation in A431 cells. Therefore, our results indicate that the 5-LOX/LTC4 /CysLT1 signaling pathway regulates EGF-induced cell migration by increasing Tiam1 expression, leading to a second wave of Rac1 activation. Thus, CysLT1 may serve as a new molecular target for antimetastatic therapy. In addition, the CysLT1 antagonist, montelukast, which is used clinically for allergy treatment, might have great potential as a novel type of antimetastatic agent. © 2013 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

  12. Differential Sulfation Remodelling of Heparan Sulfate by Extracellular 6-O-sulfatases Regulates Fibroblast Growth Factor-induced Boundary Formation By Glial Cells: Implications for Glial Cell Transplantation

    PubMed Central

    Higginson, Jennifer R; Thompson, Sophie M; Santos-Silva, Alessandra; Guimond, Scott E; Turnbull, Jeremy E; Barnett, Susan C

    2012-01-01

    Previously, it has been shown that rat Schwann cells (SCs), but not olfactory ensheathing cells (OECs), form a boundary with astrocytes, due to a SC-specific secreted factor. Here, we identify highly sulfated heparan sulfates (HS), and fibroblast growth factors (FGF) 1 and FGF9, as possible determinants of boundary formation induced by rat SCs. Disaccharide analysis of HS in SC and rat OEC conditioned medium showed that SCs secrete more highly sulfated HS than OECs. The dependence of the boundary-forming activity on high levels of sulfation was confirmed using a panel of semi-synthetic modified heparins with variable levels of sulfation. Furthermore, extracellular HS 6-O-endosulfatase enzymes, Sulf 1 and Sulf 2, were expressed at a significantly lower level by SCs compared to OECs and siRNA reduction of Sulfs in OECs was, in itself, sufficient to induce boundary formation. This demonstrates a key role for remodelling (reduction) of HS 6-O-sulfation by OECs to suppress boundary formation, in comparison to SCs. Furthermore, specific anti-FGF1 and FGF9 antibodies disrupted SC/astrocyte boundary formation, supporting a role for an HS sulfation-dependent FGF signalling mechanism via FGF receptors (FGFR) on astrocytes. We propose a model in which FGF1 and FGF9 signalling is differentially modulated by patterns of glial cell HS sulfation, dependent on Sulf 1 and Sulf 2 expression, to control FGFR3-IIIb mediated astrocytic responses. Moreover, these data suggest manipulation of HS sulfation after CNS injury as a potential novel approach for therapeutic intervention in CNS repair. PMID:23136428

  13. Combined Treatment with Troglitazone and Lovastatin Inhibited Epidermal Growth Factor-Induced Migration through the Downregulation of Cysteine-Rich Protein 61 in Human Anaplastic Thyroid Cancer Cells

    PubMed Central

    Chin, Li-Han; Hsu, Sung-Po

    2015-01-01

    Our previous studies have demonstrated that epidermal growth factor (EGF) can induce cell migration through the induction of cysteine-rich protein 61 (Cyr61) in human anaplastic thyroid cancer (ATC) cells. The aim of the present study was to determine the inhibitory effects of combined treatment with the peroxisome proliferator-activated receptor-γ (PPARγ) ligand troglitazone and the cholesterol-lowering drug lovastatin at clinically achievable concentrations on ATC cell migration. Combined treatment with 5 μM troglitazone and 1 μM lovastatin exhibited no cytotoxicity but significantly inhibited EGF-induced migration, as determined using wound healing and Boyden chamber assays. Cotreatment with troglitazone and lovastatin altered the epithelial-to-mesenchymal-transition (EMT) -related marker gene expression of the cells; specifically, E-cadherin expression increased and vimentin expression decreased. In addition, cotreatment reduced the number of filopodia, which are believed to be involved in migration, and significantly inhibited EGF-induced Cyr61 mRNA and protein expression as well as Cyr61 secretion. Moreover, the phosphorylation levels of 2 crucial signal molecules for EGF-induced Cyr61 expression, the cAMP response element-binding protein (CREB) and extracellular signal-regulated kinase (ERK), were decreased in cells cotreated with troglitazone and lovastatin. Performing a transient transfection assay revealed that the combined treatment significantly suppressed Cyr61 promoter activity. These results suggest that combined treatment with low doses of troglitazone and lovastatin effectively inhibits ATC cell migration and may serve as a novel therapeutic strategy for metastatic ATC. PMID:25742642

  14. Combined treatment with troglitazone and lovastatin inhibited epidermal growth factor-induced migration through the downregulation of cysteine-rich protein 61 in human anaplastic thyroid cancer cells.

    PubMed

    Chin, Li-Han; Hsu, Sung-Po; Zhong, Wen-Bin; Liang, Yu-Chih

    2015-01-01

    Our previous studies have demonstrated that epidermal growth factor (EGF) can induce cell migration through the induction of cysteine-rich protein 61 (Cyr61) in human anaplastic thyroid cancer (ATC) cells. The aim of the present study was to determine the inhibitory effects of combined treatment with the peroxisome proliferator-activated receptor-γ (PPARγ) ligand troglitazone and the cholesterol-lowering drug lovastatin at clinically achievable concentrations on ATC cell migration. Combined treatment with 5 μM troglitazone and 1 μM lovastatin exhibited no cytotoxicity but significantly inhibited EGF-induced migration, as determined using wound healing and Boyden chamber assays. Cotreatment with troglitazone and lovastatin altered the epithelial-to-mesenchymal-transition (EMT) -related marker gene expression of the cells; specifically, E-cadherin expression increased and vimentin expression decreased. In addition, cotreatment reduced the number of filopodia, which are believed to be involved in migration, and significantly inhibited EGF-induced Cyr61 mRNA and protein expression as well as Cyr61 secretion. Moreover, the phosphorylation levels of 2 crucial signal molecules for EGF-induced Cyr61 expression, the cAMP response element-binding protein (CREB) and extracellular signal-regulated kinase (ERK), were decreased in cells cotreated with troglitazone and lovastatin. Performing a transient transfection assay revealed that the combined treatment significantly suppressed Cyr61 promoter activity. These results suggest that combined treatment with low doses of troglitazone and lovastatin effectively inhibits ATC cell migration and may serve as a novel therapeutic strategy for metastatic ATC.

  15. Efficacy and safety of nerve growth factor for the treatment of neurological diseases: a meta-analysis of 64 randomized controlled trials involving 6,297 patients

    PubMed Central

    Zhao, Meng; Li, Xiao-yan; Xu, Chun-ying; Zou, Li-ping

    2015-01-01

    OBJECTIVE: China is the only country where nerve growth factor is approved for large-scale use as a clinical medicine. More than 10 years ago, in 2003, nerve growth factor injection was listed as a national drug. The goal of this article is to evaluate comprehensively the efficacy and safety of nerve growth factor for the treatment of neurological diseases. DATA RETRIEVAL: A computer-based retrieval was performed from six databases, including the Cochrane Library, PubMed, EMBASE, Sino Med, CNKI, and the VIP database, searching from the clinical establishment of nerve growth factor for treatment until December 31, 2013. The key words for the searches were “nerve growth factor, randomized controlled trials” in Chinese and in English. DATA SELECTION: Inclusion criteria: any study published in English or Chinese referring to randomized controlled trials of nerve growth factor; patients with neurological diseases such as peripheral nerve injury, central nerve injury, cranial neuropathy, and nervous system infections; patients older than 7 years; similar research methods and outcomes assessing symptoms; and measurement of nerve conduction velocities. The meta-analysis was conducted using Review Manager 5.2.3 software. MAIN OUTCOME MEASURES: The total effective rate, the incidence of adverse effects, and the nerve conduction velocity were recorded for each study. RESULTS: Sixty-four studies involving 6,297 patients with neurological diseases were included. The total effective rate in the group treated with nerve growth factor was significantly higher than that in the control group (P < 0.0001, RR: 1.35, 95%CI: 1.30–1.40). The average nerve conduction velocity in the nerve growth factor group was significantly higher than that in the control group (P < 0.00001, MD: 4.59 m/s, 95%CI: 4.12–5.06). The incidence of pain or scleroma at the injection site in the nerve growth factor group was also higher than that in the control group (P < 0.00001, RR: 6.30, 95%CI: 3.53

  16. Type II cGMP-dependent protein kinase inhibits epidermal growth factor-induced phosphatidylinositol-3-kinase/Akt signal transduction in gastric cancer cells.

    PubMed

    Wu, Min; Chen, Yongchang; Jiang, Lu; Li, Yueying; Lan, Ting; Wang, Ying; Qian, Hai

    2013-12-01

    Our previous study revealed that Type II cGMP-dependent protein kinase (PKG II) inhibits epidermal growth factor (EGF)-induced MAPK/ERK and MAPK/JNK-mediated signal transduction through the inhibition of the phosphorylation/activation of the EGF receptor (EGFR). As EGFR also mediates several other signal transduction pathways besides MAPK-mediated pathways, the present study was designed to investigate whether PKG II was able to inhibit EGF/EGFR-induced phosphatidylinositol-3-kinase (PI3K)/Akt-mediated signal transduction. The AGS human gastric cancer cell line was infected with adenoviral constructs encoding a cDNA of PKG II (Ad-PKG II) to increase the expression of PKG II, and treated with 8-pCPT-cGMP to activate the enzyme. Western blotting was used to detect the phosphorylation/activation of the key components of the signal transduction pathway, including EGFR, PI3K, Akt, mTOR and NF-κB. The levels of apoptosis-related proteins, including Bax, Bcl-2, caspase 9 and DNA fragment factor (DFF), were also determined by western blotting. Terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling staining was used to detect the apoptosis of the AGS cells. The results revealed that EGF treatment increased the phosphorylation (activation) of EGFR, PI3K, Akt and mTOR, and increased the nuclear localization (activation) of NF-κB. EGF treatment also reduced the apoptosis of the AGS cells and increased the expression of the anti-apoptotic protein, Bcl-2, but had no effect on the expression of the pro-apoptotic protein, Bax, and did not alter the levels of caspase 9 and DFF. Increasing the PKG II activity of AGS cells by infecting them with Ad-PKG II and stimulating them with 8-pCPT-cGMP inhibited the EGF-induced activation of EGFR, PI3K, Akt, mTOR and NF-κB; caused an increase in caspase 9 breakdown (activation) and DFF levels; and reversed the anti-apoptotic effect of EGF. The results suggest that PKG II may also inhibit EGF-induced signal transduction of PI3

  17. Zedoarondiol Inhibits Platelet-Derived Growth Factor-Induced Vascular Smooth Muscle Cells Proliferation via Regulating AMP-Activated Protein Kinase Signaling Pathway.

    PubMed

    Mao, Huimin; Tao, Tianqi; Song, Dandan; Liu, Mi; Wang, Xiaoren; Liu, Xiuhua; Shi, Dazhuo

    2016-01-01

    Vascular smooth muscle cells (VSMCs) proliferation contributes significantly to atherosclerosis and in-stent restenosis. Platelet-derived growth factor-BB (PDGF-BB) plays a vital role in VSMCs proliferation. Zedoarondiol, a sesquiterpene lactone compound, has an anti-inflammatory activity. However, the role of zedoarondiol in PDGF-BB-mediated VSMCs proliferation remains unclear. In this study, we investigated the effects of zedoarondiol on PDGF-BB-induced VSMCs proliferation and explored the possible mechanisms. The inhibitory effects of zedoarondiol on PDGF-BB-induced VSMCs proliferation were evaluated by direct cell counting and the Cell Counting Kit-8 (CCK-8) assay. DNA synthesis was examined by bromodeoxyuridine (BrdU) incorporation assay. Cell cycle was assessed by propidium iodide staining. Western blotting was performed to determine the expression of cyclin-dependent kinase 2 (CDK2), cyclin E, p53, p21, total and phosphorylated adenosine monophosphate-activated protein kinase (AMPK), acetyl CoA carboxylase (ACC), mammalian target of rapamycin (mTOR), and p70 ribosomal protein S6 kinase (p70S6K). Zedoarondiol suppressed PDGF-BB-induced VSMCs proliferation and DNA synthesis, and induced cell cycle arrest in G0/G1 phase. In addition, zedoarondiol activated AMPK and ACC, inhibited the phosphorylation of mTOR and p70S6K, increased the expression of p53 and p21, and decreased the expression of CDK2 and cyclin E. Compound C (an AMPK inhibitor) abrogated, whereas 5-aminoimidazole-4-carboxamide 1-β-ribofuranoside (AICAR, an AMPK activator) enhanced zedoarondiol-mediated inhibition of VSMCs proliferation and DNA synthesis. Zedoarondiol inhibits PDGF-BB-induced VSMCs proliferation via AMPK-mediated down-regulation of the mTOR/p70S6K pathway and up-regulation of the p53/p21 pathway. These findings suggest that zedoarondiol might be a promising compound against atherosclerosis and in-stent restenosis. © 2016 The Author(s) Published by S. Karger AG, Basel.

  18. Therapeutic effect of nerve growth factor on cerebral infarction in dogs using the hemisphere anomalous volume ratio of diffusion-weighted magnetic resonance imaging.

    PubMed

    Wang, Yong; Zhang, Hui; Wang, Zhe; Geng, Zuojun; Liu, Huaijun; Yang, Haiqing; Song, Peng; Liu, Qing

    2012-08-25

    A model of focal cerebral ischemic infarction was established in dogs through middle cerebral artery occlusion of the right side. Thirty minutes after occlusion, models were injected with nerve growth factor adjacent to the infarct locus. The therapeutic effect of nerve growth factor against cerebral infarction was assessed using the hemisphere anomalous volume ratio, a quantitative index of diffusion-weighted MRI. At 6 hours, 24 hours, 7 days and 3 months after modeling, the hemisphere anomalous volume ratio was significantly reduced after treatment with nerve growth factor. Hematoxylin-eosin staining, immunohistochemistry, electron microscopy and neurological function scores showed that infarct defects were slightly reduced and neurological function significantly improved after nerve growth factor treatment. This result was consistent with diffusion-weighted MRI measurements. Experimental findings indicate that nerve growth factor can protect against cerebral infarction, and that the hemisphere anomalous volume ratio of diffusion-weighted MRI can be used to evaluate the therapeutic effect.

  19. Therapeutic effect of nerve growth factor on cerebral infarction in dogs using the hemisphere anomalous volume ratio of diffusion-weighted magnetic resonance imaging★

    PubMed Central

    Wang, Yong; Zhang, Hui; Wang, Zhe; Geng, Zuojun; Liu, Huaijun; Yang, Haiqing; Song, Peng; Liu, Qing

    2012-01-01

    A model of focal cerebral ischemic infarction was established in dogs through middle cerebral artery occlusion of the right side. Thirty minutes after occlusion, models were injected with nerve growth factor adjacent to the infarct locus. The therapeutic effect of nerve growth factor against cerebral infarction was assessed using the hemisphere anomalous volume ratio, a quantitative index of diffusion-weighted MRI. At 6 hours, 24 hours, 7 days and 3 months after modeling, the hemisphere anomalous volume ratio was significantly reduced after treatment with nerve growth factor. Hematoxylin-eosin staining, immunohistochemistry, electron microscopy and neurological function scores showed that infarct defects were slightly reduced and neurological function significantly improved after nerve growth factor treatment. This result was consistent with diffusion-weighted MRI measurements. Experimental findings indicate that nerve growth factor can protect against cerebral infarction, and that the hemisphere anomalous volume ratio of diffusion-weighted MRI can be used to evaluate the therapeutic effect. PMID:25624813

  20. Epidermal growth factor-induced selective phosphorylation of cultured rat hepatocyte 55-kD cytokeratin before filament reorganization and DNA synthesis

    PubMed Central

    1989-01-01

    We have reported previously that the addition of dexamethasone to cultured quiescent suckling rat hepatocytes in the presence of insulin, a culture condition which does not cause growth activation, induces a selective increase in the synthesis of the 49-kD/55-kD cytokeratin (CK49/CK55) pair over a 24-h period. This increased synthesis coincides with the formation of dense filament networks reminiscent of those observed in situ at the cell periphery (Marceau, N., H. Baribault, and I. Leroux-Nicollet. 1985. Can. J. Biochem. Cell Biol. 63:448-457). We show here for the first time that when EGF is added 48 h after insulin and dexamethasone, there is an early preferential phosphorylation of the CK55 of the CK49/CK55 pair, an induced filament rearrangement from the cell periphery to the cytoplasm, and a subsequent entry into S phase and mitosis after a lag period of 8 h. Indirect immunofluorescence microscopy with monoclonal antibodies to CK49 and CK55 indicate that, while before EGF treatment the cytokeratin filaments were mainly distributed near the cell periphery, the addition of EGF resulted in their reorganization to a predominantly cytoplasmic localization within less than 3 h. Antitubulin and anti-actin antibodies showed no detectable alteration in the distribution of microtubules and microfilaments. Pulse-chase measurements with [35S]methionine showed no apparent change in the turnover of either CK49 or CK55 during the period that precedes the initiation of DNA synthesis. 32P-labeling in vivo followed by SDS-PAGE demonstrated that CK55 was phosphorylated at a much higher level than CK49 in nonstimulated hepatocytes, and that the addition of EGF resulted in a selective stimulation of 32P-CK55 labeling within less than 30 min. Comparative analyses by two-dimensional PAGE of [35S]methionine and 32P- labeled cytokeratins at various times after EGF stimulation demonstrated a rapid increase in a first phosphorylated form of CK55 and the appearance of a second

  1. Adiponectin Is Involved in Connective Tissue Growth Factor-Induced Proliferation, Migration and Overproduction of the Extracellular Matrix in Keloid Fibroblasts.

    PubMed

    Luo, Limin; Li, Jun; Liu, Han; Jian, Xiaoqing; Zou, Qianlei; Zhao, Qing; Le, Qu; Chen, Hongdou; Gao, Xinghua; He, Chundi

    2017-05-12

    Adiponectin, an adipocyte-derived hormone, exerts pleiotropic biological effects on metabolism, inflammation, vascular homeostasis, apoptosis and immunity. Recently, adiponectin has been suggested to attenuate the progression of human dermal fibrosis. Connective tissue growth factor (CTGF) is induced in keloids and is thought to be participated in the formation of keloid fibrosis. However, the roles played by adiponectin in keloids remain unclear. In this study, we explored the effects of adiponectin on CTGF-induced cell proliferation, migration and the deposition of extracellular matrix (ECM) and their associated intracellular signalling pathways in keloid fibroblasts (KFs). We also explored possible mechanisms of keloid pathogenesis. Primary fibroblast cultures were established from foreskin biopsies and skin biopsies from patients with keloids. The expression of adiponectin and adiponectin receptors (adipoRs) was evaluated by reverse transcription-PCR (RT-PCR), quantitative real-time RT-PCR, immunofluorescence staining, and immunohistochemical analysis. Next, KFs and normal dermal fibroblasts (NFs) were treated with CTGF in the presence or absence of adiponectin. A cell counting kit-8 (CCK-8) and the Transwell assay were used to examine cell proliferation and migration. The level of the collagen I, fibronectin (FN) and α-smooth muscle actin (α-SMA) mRNAs and proteins were determined by quantitative real-time RT-PCR and western blotting. The effects of RNA interference (RNAi) targeting the adipoR genes were detected. Phosphorylation of adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK), mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3 kinase-protein kinase (PI3K-Akt) were examined by western blotting to further investigate the signalling pathways. Furthermore, inhibitors of signal transduction pathways were investigated. The expression levels of adiponectin and adipoRs were significantly decreased in keloids compared with those

  2. Phosphorylation of p85 beta PIX, a Rac/Cdc42-specific guanine nucleotide exchange factor, via the Ras/ERK/PAK2 pathway is required for basic fibroblast growth factor-induced neurite outgrowth.

    PubMed

    Shin, Eun-Young; Shin, Kyung-Sun; Lee, Chan-Soo; Woo, Kyung-Nam; Quan, Song-Hua; Soung, Nak-Kyun; Kim, Young Gyu; Cha, Choong Ik; Kim, Seung-Ryul; Park, Dongeun; Bokoch, Gary M; Kim, Eung-Gook

    2002-11-15

    Guanine nucleotide exchange factors (GEFs) have been implicated in growth factor-induced neuronal differentiation through the activation of small GTPases. Although phosphorylation of these GEFs is considered an activation mechanism, little is known about the upstream of PAK-interacting exchange factor (PIX), a member of the Dbl family of GEFs. We report here that phosphorylation of p85 betaPIX/Cool/p85SPR is mediated via the Ras/ERK/PAK2 pathway. To understand the role of p85 betaPIX in basic fibroblast growth factor (bFGF)-induced neurite outgrowth, we established PC12 cell lines that overexpress the fibroblast growth factor receptor-1 in a tetracycline-inducible manner. Treatment with bFGF induces the phosphorylation of p85 betaPIX, as determined by metabolic labeling and mobility shift upon gel electrophoresis. Interestingly, phosphorylation of p85 betaPIX is inhibited by PD98059, a specific MEK inhibitor, suggesting the involvement of the ERK cascade. PAK2, a major PAK isoform in PC12 cells as well as a binding partner of p85 betaPIX, also functions upstream of p85 betaPIX phosphorylation. Surprisingly, PAK2 directly binds to ERK, and its activation is dependent on ERK. p85 betaPIX specifically localizes to the lamellipodia at neuronal growth cones in response to bFGF. A mutant form of p85 betaPIX (S525A/T526A), in which the major phosphorylation sites are replaced by alanine, shows significant defect in targeting. Moreover, expression of the mutant p85 betaPIX efficiently blocks PC12 cell neurite outgrowth. Our study defines a novel signaling pathway for bFGF-induced neurite outgrowth that involves activation of the PAK2-p85 betaPIX complex via the ERK cascade and subsequent translocation of this complex.

  3. Expression of nerve growth factor and its receptors in the uterus of rabbits: functional involvement in prostaglandin synthesis.

    PubMed

    Maranesi, M; Parillo, F; Leonardi, L; Rebollar, P G; Alonso, B; Petrucci, L; Gobbetti, A; Boiti, C; Arruda-Alencar, J; Moura, A; Zerani, M

    2016-07-01

    The aim of the present study was to evaluate: (1) the presence of nerve growth factor (NGF), neurotrophic tyrosine kinase receptor 1 (NTRK1), and nerve growth factor receptor (NGFR) in the rabbit uterus; and (2) the in vitro effects of NGF on PGF2α and PGE2 synthesis and on the PGE2-9-ketoreductase (PGE2-9-K) activity by the rabbit uterus. Nerve growth factor, NTRK1, and NGFR were immunolocalized in the luminal and glandular epithelium and stroma cells of the endometrium. reverse transcriptase polymerase chain reaction indicated the presence of messenger RNA for NGF, NTRK1, and NGFR in the uterus. Nerve growth factor increased (P < 0.01) in vitro secretions of PGF2α and PGE2 but coincubation with either NTRK1 or oxide nitric synthase (NOS) inhibitors reduced (P < 0.01) PGF2α production and blocked (P < 0.01) PGE2 secretion. Prostaglandins releases were lower (P < 0.01) than control when uterine samples were treated with NGF plus cyclooxygenase inhibitor. However, addition of NGFR inhibitor reduced (P < 0.01) PGF2α secretion less efficiently than NTRK1 or NOS inhibitors but had no effect on PGE2 yield. Nerve growth factor increased (P < 0.01) the activity of PGE2-9-K, whereas coincubation with NTRK1 or NOS inhibitors abolished (P < 0.01) this increase in PGE2-9-K activity. However, cotreatment with either cyclooxygenase or NGFR inhibitors had no effect on PGE2-9-K activity. This is the first study to document the distribution of NGF/NTRK1 and NGFR systems and their effects on prostaglandin synthesis in the rabbit uterus. NGF/NTRK1 increases PGF2α and PGE2 productions by upregulating NOS and PGE2-9-K activities, whereas NGF/NGFR augments only PGF2α secretion, through an intracellular mechanism that is still unknown.

  4. Hypertrophy of neurons within cardiac ganglia in human, canine, and rat heart failure: the potential role of nerve growth factor.

    PubMed

    Singh, Sanjay; Sayers, Scott; Walter, James S; Thomas, Donald; Dieter, Robert S; Nee, Lisa M; Wurster, Robert D

    2013-08-19

    Autonomic imbalances including parasympathetic withdrawal and sympathetic overactivity are cardinal features of heart failure regardless of etiology; however, mechanisms underlying these imbalances remain unknown. Animal model studies of heart and visceral organ hypertrophy predict that nerve growth factor levels should be elevated in heart failure; whether this is so in human heart failure, though, remains unclear. We tested the hypotheses that neurons in cardiac ganglia are hypertrophied in human, canine, and rat heart failure and that nerve growth factor, which we hypothesize is elevated in the failing heart, contributes to this neuronal hypertrophy. Somal morphology of neurons from human (579.54±14.34 versus 327.45±9.17 μm(2); P<0.01) and canine hearts (767.80±18.37 versus 650.23±9.84 μm(2); P<0.01) failing secondary to ischemia and neurons from spontaneously hypertensive rat hearts (327.98±3.15 versus 271.29±2.79 μm(2); P<0.01) failing secondary to hypertension reveal significant hypertrophy of neurons in cardiac ganglia compared with controls. Western blot analysis shows that nerve growth factor levels in the explanted, failing human heart are 250% greater than levels in healthy donor hearts. Neurons from cardiac ganglia cultured with nerve growth factor are significantly larger and have greater dendritic arborization than neurons in control cultures. Hypertrophied neurons are significantly less excitable than smaller ones; thus, hypertrophy of vagal postganglionic neurons in cardiac ganglia would help to explain the parasympathetic withdrawal that accompanies heart failure. Furthermore, our observations suggest that nerve growth factor, which is elevated in the failing human heart, causes hypertrophy of neurons in cardiac ganglia.

  5. Pituitary adenylyl cyclase-activating polypeptide and nerve growth factor use the proteasome to rescue nerve growth factor-deprived sympathetic neurons cultured from chick embryos.

    PubMed

    Przywara, D A; Kulkarni, J S; Wakade, T D; Leontiev, D V; Wakade, A R

    1998-11-01

    Removal of nerve growth factor (NGF) from sympathetic neurons initiates a neuronal death program and apoptosis. We show that pituitary adenylyl cyclase-activating polypeptide (PACAP) prevents apoptosis in NGF-deprived sympathetic neurons. PACAP (100 nM) added to culture medium at the time of plating failed to support neuronal survival. However, in neurons grown for 2 days with NGF and then deprived of NGF, PACAP prevented cell death for the next 24-48 h. Uptake of [3H]norepinephrine ([3H]NE) was used as an index of survival and decreased >50% in NGF-deprived cultures within 24 h. PACAP (1-100 nM) restored [3H]NE uptake to 92 +/- 8% of that of NGF-supported controls. Depolarization-induced [3H]NE release in neurons rescued by PACAP was the same as that in NGF-supported neurons. PACAP rescue was not mimicked by forskolin or 8-bromo-cyclic AMP and was not blocked by the protein kinase A inhibitor Rp-adenosine 3',5'-cyclic monophosphothioate. Mobilization of phosphatidylinositol by muscarine failed to support NGF-deprived neurons. Thus, PACAP may use novel signaling to promote survival of sympathetic neurons. The apoptosis-associated caspase CPP32 activity increased approximately fourfold during 6 h of NGF withdrawal (145 +/- 40 versus 38 +/- 17 nmol of substrate cleaved/min/mg of protein) and returned to even below the control level in NGF-deprived, PACAP-rescued cultures (14 +/- 7 nmol/min/mg of protein). Readdition of NGF or PACAP to NGF-deprived cultures reversed CPP32 activation, and this was blocked by lactacystin, a potent and specific inhibitor of the 20S proteasome, suggesting that NGF and PACAP target CPP32 for destruction by the proteasome. As PACAP is a preganglionic neurotransmitter in autonomic ganglia, we propose a novel function for this transmitter as an apoptotic rescuer of sympathetic neurons when the supply of NGF is compromised.

  6. Expression of nerve growth factor and neurotrophin-3 mRNAs in hippocampal interneurons: morphological characterization, levels of expression, and colocalization of nerve growth factor and neurotrophin-3.

    PubMed

    Pascual, M; Rocamora, N; Acsády, L; Freund, T F; Soriano, E

    1998-05-25

    We have investigated the distribution and morphology of hippocampal interneurons that express the neurotrophins nerve growth factor (NGF) and neurotrophin-3 (NT-3) in the rat. For this study, we combined in situ hybridization for the detection of NGF and NT-3 mRNAs and immunocytochemistry against the calcium-binding proteins parvalbumin (PARV), calretinin (CALR), and calbindin (CALB). Whereas the majority of PARV+ interneurons expressed NGF mRNA, only subsets of CALR- and CALB-immunoreactive interneurons (23% and 24%, respectively) displayed NGF hybridization. Most CALB/NGF+ cells were located in the stratum oriens/alveus of the CA3-CA1 regions, suggesting that they may include the population of CALB+, hippocamposeptal, nonpyramidal neurons. Most of the nonspiny CALR/NGF+ neurons were located within or in the vicinity of the pyramidal layer and had faint CALR immunostaining and stellate, thin dendrites. Regarding the spiny CALR-immunoreactive cells, we found that most of these neurons in the hilus were NGF+, whereas only 59% of displayed NGF hybridization in the stratum lucidum of the CA3 region. A small subset of PARV- and CALR-immunoreactive cells expressed NT-3 mRNA (16% and 13%, respectively). NT-3 message was not found in the large basket cells of the dentate gyrus, whereas the distribution and morphology of CALR/NT-3+ cells were similar to those of nonspiny CALR/NGF+ cells. In fact, double in situ hybridization analysis confirmed that most NT-3+ neurons also expressed NGF mRNA, indicating coexpression of both neurotrophins in subpopulations of PARV+ and CALR+ neurons. Moreover, the level of NGF mRNA expression was higher in PARV+ neurons than in CALR- and CALB-immunoreactive interneurons, whereas NT-3 message was expressed similarly in PARV+ and CALR+ neurons. The present findings show a differential expression of NGF and NT-3 mRNAs in subsets of hippocampal interneurons and suggest that the expression of these transcripts depends on factors intrinsic to

  7. Exploring Serum Levels of Brain Derived Neurotrophic Factor and Nerve Growth Factor Across Glaucoma Stages

    PubMed Central

    Busanello, Anna; Bonini, Stefano; Quaranta, Luciano; Agnifili, Luca; Manni, Gianluca

    2017-01-01

    Purpose To investigate the serum levels of Brain Derived Neurotrophic Factor (BDNF) and Nerve Growth Factor (NGF) in patients affected by primary open angle glaucoma with a wide spectrum of disease severity compared to healthy controls and to explore their relationship with morphological and functional glaucoma parameters. Materials and Methods 45 patients affected by glaucoma at different stages and 15 age-matched healthy control subjects underwent visual field testing, peripapillary retinal nerve fibre layer thickness measurement using Spectral Domain Optical Coherence Tomography and blood collection for both neurotrophins detection by Enzyme-Linked Immunosorbent Assay. Statistical analysis and association between biostrumental and biochemical data were investigated. Results Serum levels of BDNF in glaucoma patients were significantly lower than those measured in healthy controls (261.2±75.0 pg/ml vs 313.6±79.6 pg/ml, p = 0.03). Subgroups analysis showed that serum levels of BDNF were significantly lower in early (253.8±40.7 pg/ml, p = 0.019) and moderate glaucoma (231.3±54.3 pg/ml, p = 0.04) but not in advanced glaucoma (296.2±103.1 pg/ml, p = 0.06) compared to healthy controls. Serum levels of NGF in glaucoma patients were significantly lower than those measured in the healthy controls (4.1±1 pg/mL vs 5.5±1.2 pg/mL, p = 0.01). Subgroups analysis showed that serum levels of NGF were significantly lower in early (3.5±0.9 pg/mL, p = 0.0008) and moderate glaucoma (3.8±0.7 pg/ml, p<0.0001) but not in advanced glaucoma (5.0±0.7 pg/ml, p = 0.32) compared to healthy controls. BDNF serum levels were not related to age, visual field mean deviation or retinal nerve fibre layer thickness either in glaucoma or in controls while NGF levels were significantly related to visual field mean deviation in the glaucoma group (r2 = 0.26, p = 0.004). Conclusions BDNF and NGF serum levels are reduced in the early and moderate glaucoma stages, suggesting the possibility that

  8. Uptake of nerve growth factor along peripheral and spinal axons of primary sensory neurons

    SciTech Connect

    Richardson, P.M.; Riopelle, R.J.

    1984-07-01

    To investigate the distribution of nerve growth factor (NGF) receptors on peripheral and central axons, (/sup 125/I)NGF was injected into the sciatic nerve or spinal cord of adult rats. Accumulation of (/sup 125/I)NGF in lumbar dorsal root ganglia was monitored by gamma emission counting and radioautography. (/sup 125/I)NGF, injected endoneurially in small quantities, was taken into sensory axons by a saturable process and was transported retrogradely to their cell bodies at a maximal rate of 2.5 to 7.5 mm/hr. Because very little (/sup 125/I)NGF reached peripheral terminals, the results were interpreted to indicate that receptors for NGF are present on nonterminal segments of sensory axons. The specificity and high affinity of NGF uptake were illustrated by observations that negligible amounts of gamma activity accumulated in lumbar dorsal root ganglia after comparable intraneural injection of (/sup 125/I) cytochrome C or (/sup 125/I)oxidized NGF. Similar techniques were used to demonstrate avid internalization and retrograde transport of (/sup 125/I)NGF by intraspinal axons arising from dorsal root ganglia. Following injection of (/sup 125/I)NGF into lumbar or cervical regions of the spinal cord, neuronal perikarya were clearly labeled in radioautographs of lumbar dorsal root ganglia. Sites for NGF uptake on primary sensory neurons in the adult rat are not restricted to peripheral axon terminals but are extensively distributed along both peripheral and central axons. Receptors on axons provide a mechanism whereby NGF supplied by glia could influence neuronal maintenance or axonal regeneration.

  9. A nerve growth factor-regulated messenger RNA encodes a new intermediate filament protein

    PubMed Central

    1988-01-01

    Differential screening of a cDNA library from the PC12 rat pheochromocytoma cell line previously revealed a clone, clone 73, whose corresponding mRNA is induced by nerve growth factor (NGF). Induction parallels NGF-stimulated PC12 differentiation from a chromaffinlike phenotype to a sympathetic neuronlike phenotype. We report that DNA sequence analysis reveals that clone 73 mRNA encodes an intermediate filament (IF) protein whose predicted amino acid sequence is distinct from the known sequences of other members of the IF protein family. The sequence has highest homology with desmin and vimentin and includes the highly conserved central alpha-helical rod domain with the characteristic heptad repeat of hydrophobic residues, but has lower homology in the amino-terminal head and carboxyl-terminal tail domains. The head domain contains a large number of serine residues which are potential phosphorylation sites. The expression of clone 73 in vivo in the nervous system of the adult rat was investigated by in situ hybridization of clone 73 probes to tissue sections. The mRNA is expressed at high levels in ganglia of the peripheral nervous system, including the superior cervical ganglion (sympathetic), ciliary ganglion (parasympathetic), and dorsal root ganglion (sensory). In the central nervous system, motor nuclei of cranial nerves III, IV, V, VI, VII, X, and XII as well as ventral horn motor neurons and a restricted set of other central nervous system nuclei express the clone 73 mRNA. Tissues apart from those of the nervous system did not in general express the mRNA, with only very low levels detected in adrenal gland. We discuss the implications of these results for the mechanism of NGF- induced PC12 cell differentiation, the pathways of neuronal development in vivo, and the possible function of the clone 73 IF protein and its relationship to other IF proteins. PMID:3339087

  10. Tamoxifen inhibits malignant peripheral nerve sheath tumor growth in an estrogen receptor–independent manner

    PubMed Central

    Byer, Stephanie J.; Eckert, Jenell M.; Brossier, Nicole M.; Clodfelder-Miller, Buffie J.; Turk, Amy N.; Carroll, Andrew J.; Kappes, John C.; Zinn, Kurt R.; Prasain, Jeevan K.; Carroll, Steven L.

    2011-01-01

    Few therapeutic options are available for malignant peripheral nerve sheath tumors (MPNSTs), the most common malignancy associated with neurofibromatosis type 1 (NF1). Guided by clinical observations suggesting that some NF1-associated nerve sheath tumors are hormonally responsive, we hypothesized that the selective estrogen receptor (ER) modulator tamoxifen would inhibit MPNST tumorigenesis in vitro and in vivo. To test this hypothesis, we examined tamoxifen effects on MPNST cell proliferation and survival, MPNST xenograft growth, and the mechanism by which tamoxifen impeded these processes. We found that 1–5 μM 4-hydroxy-tamoxifen induced MPNST cell death, whereas 0.01–0.1 μM 4-hydroxy-tamoxifen inhibited mitogenesis. Dermal and plexiform neurofibromas, MPNSTs, and MPNST cell lines expressed ERβ and G-protein-coupled ER-1 (GPER); MPNSTs also expressed estrogen biosynthetic enzymes. However, MPNST cells did not secrete 17β-estradiol, exogenous 17β-estradiol did not stimulate mitogenesis or rescue 4-hydroxy-tamoxifen effects on MPNST cells, and the steroidal antiestrogen ICI-182,780 did not mimic tamoxifen effects on MPNST cells. Further, ablation of ERβ and GPER had no effect on MPNST proliferation, survival, or tamoxifen sensitivity, indicating that tamoxifen acts via an ER-independent mechanism. Consistent with this hypothesis, inhibitors of calmodulin (trifluoperazine, W-7), another known tamoxifen target, recapitulated 4-hydroxy-tamoxifen effects on MPNST cells. Tamoxifen was also effective in vivo, demonstrating potent antitumor activity in mice orthotopically xenografted with human MPNST cells. We conclude that 4-hydroxy-tamoxifen inhibits MPNST cell proliferation and survival via an ER-independent mechanism. The in vivo effectiveness of tamoxifen provides a rationale for clinical trials in cases of MPNSTs. PMID:21075781

  11. Nerve growth factor stimulates cellular proliferation of human epithelial ovarian cancer.

    PubMed

    Urzua, U; Tapia, V; Geraldo, M P; Selman, A; Vega, M; Romero, C

    2012-09-01

    Due to its ability to induce vascular endothelial growth factor expression and proliferation, migration, and vasculogenesis of endothelial cells, nerve growth factor (NGF) has been considered as an angiogenic factor in epithelial ovarian cancer (EOC). In this work, we evaluated the angiogenic and proliferative mRNA expression profiles of EOC and addressed the responsiveness of EOC explants to NGF stimulation. Twenty EOC samples were obtained from Obstetrics and Gynecology Department, University of Chile's Clinical Hospital. Global gene expression profiles of selected poorly differentiated serous EOC samples were obtained with DNA oligonucleotide microarrays. In addition, EOC explants were subjected to NGF stimulation and levels of p-AKT, BAX, BCL2, Ki-67, c-MYC, and FOXL2 proteins were determined by immunohistochemistry. Results showed that mRNAs coding for specific transcriptional regulators and antiapoptotic components of the NGF signaling pathway were upregulated in EOC cells. At the protein level, key members of the NGF pathway including p-AKT, BCL2/BAX, Ki-67, and c-MYC were found increased, while FOXL2 was decreased in response to NGF stimulation. These findings strongly suggest that NGF stimulates cellular proliferation of human EOC. © Georg Thieme Verlag KG Stuttgart · New York.

  12. Role of Nerve Growth Factor (NGF) and miRNAs in Epithelial Ovarian Cancer

    PubMed Central

    Retamales-Ortega, Rocío; Oróstica, Lorena; Vera, Carolina; Cuevas, Paula; Hernández, Andrea; Hurtado, Iván; Vega, Margarita; Romero, Carmen

    2017-01-01

    Ovarian cancer is the eighth most common cancer in women worldwide, and epithelial ovarian cancer (EOC) represents 90% of cases. Nerve growth factor (NGF) and its high affinity receptor tyrosine kinase A receptor (TRKA) have been associated with the development of several types of cancer, including EOC; both NGF and TRKA levels are elevated in this pathology. EOC presents high angiogenesis and several molecules have been reported to induce this process. NGF increases angiogenesis through its TRKA receptor on endothelial cells, and by indirectly inducing vascular endothelial growth factor expression. Other molecules controlled by NGF include ciclooxigenase-2, disintegrin and metalloproteinase domain-containing protein 17 (ADAM17) and calreticulin (CRT), proteins involved in crucial processes needed for EOC progression. These molecules could be modified through microRNA regulation, which could be regulated by NGF. MicroRNAs are the widest family of non-coding RNAs; they bind to 3′-UTR of mRNAs to inhibit their translation, to deadenilate or to degraded them. In EOC, a deregulation in microRNA expression has been described, including alterations of miR-200 family, cluster-17-92, and miR-23b, among others. Since the NGF-microRNA relationship in pathologies has not been studied, this review proposes that some microRNAs could be associated with NGF/TRKA activation, modifying protein levels needed for EOC progression. PMID:28245631

  13. Role of Nerve Growth Factor (NGF) and miRNAs in Epithelial Ovarian Cancer.

    PubMed

    Retamales-Ortega, Rocío; Oróstica, Lorena; Vera, Carolina; Cuevas, Paula; Hernández, Andrea; Hurtado, Iván; Vega, Margarita; Romero, Carmen

    2017-02-26

    Ovarian cancer is the eighth most common cancer in women worldwide, and epithelial ovarian cancer (EOC) represents 90% of cases. Nerve growth factor (NGF) and its high affinity receptor tyrosine kinase A receptor (TRKA) have been associated with the development of several types of cancer, including EOC; both NGF and TRKA levels are elevated in this pathology. EOC presents high angiogenesis and several molecules have been reported to induce this process. NGF increases angiogenesis through its TRKA receptor on endothelial cells, and by indirectly inducing vascular endothelial growth factor expression. Other molecules controlled by NGF include ciclooxigenase-2, disintegrin and metalloproteinase domain-containing protein 17 (ADAM17) and calreticulin (CRT), proteins involved in crucial processes needed for EOC progression. These molecules could be modified through microRNA regulation, which could be regulated by NGF. MicroRNAs are the widest family of non-coding RNAs; they bind to 3'-UTR of mRNAs to inhibit their translation, to deadenilate or to degraded them. In EOC, a deregulation in microRNA expression has been described, including alterations of miR-200 family, cluster-17-92, and miR-23b, among others. Since the NGF-microRNA relationship in pathologies has not been studied, this review proposes that some microRNAs could be associated with NGF/TRKA activation, modifying protein levels needed for EOC progression.

  14. Conformational Rigidity within Plasticity Promotes Differential Target Recognition of Nerve Growth Factor

    PubMed Central

    Paoletti, Francesca; de Chiara, Cesira; Kelly, Geoff; Covaceuszach, Sonia; Malerba, Francesca; Yan, Robert; Lamba, Doriano; Cattaneo, Antonino; Pastore, Annalisa

    2016-01-01

    Nerve Growth Factor (NGF), the prototype of the neurotrophin family, is essential for maintenance and growth of different neuronal populations. The X-ray crystal structure of NGF has been known since the early '90s and shows a β-sandwich fold with extensive loops that are involved in the interaction with its binding partners. Understanding the dynamical properties of these loops is thus important for molecular recognition. We present here a combined solution NMR/molecular dynamics study which addresses the question of whether and how much the long loops of NGF are flexible and describes the N-terminal intrinsic conformational tendency of the unbound NGF molecule. NMR titration experiments allowed identification of a previously undetected epitope of the anti-NGF antagonist antibody αD11 which will be of crucial importance for future drug lead discovery. The present study thus recapitulates all the available structural information and unveils the conformational versatility of the relatively rigid NGF loops upon functional ligand binding. PMID:28083536

  15. Association of nerve growth factor receptors with the triton X-100 cytoskeleton of PC12 cells

    SciTech Connect

    Vale, R.D.; Ignatius, M.J.; Shooter, E.M.

    1985-10-01

    Triton X-100 solubilizes membranes of PC12 cells and leaves behind a nucleus and an array of cytoskeletal filaments. Nerve growth factor (NGF) receptors are associated with this Triton X-100-insoluble residue. Two classes of NGF receptors are found on PC12 cells which display rapid and slow dissociating kinetics. Although rapidly dissociating binding is predominant (greater than 75%) in intact cells, the majority of binding to the Triton X-100 cytoskeleton is slowly dissociating (greater than 75%). Rapidly dissociating NGF binding on intact cells can be converted to a slowly dissociating form by the plant lectin wheat germ agglutinin (WGA). This lectin also increases the number of receptors which associate with the Triton X-100 cytoskeleton by more than 10-fold. SVI-NGF bound to receptors can be visualized by light microscopy autoradiography in Triton X-100-insoluble residues of cell bodies, as well as growth cones and neurites. The WGA-induced association with the cytoskeleton, however, is not specific for the NGF receptor. Concentrations of WGA which change the Triton X-100 solubility of membrane glycoproteins are similar to those required to alter the kinetic state of the NGF receptor. Both events may be related to the crossbridging of cell surface proteins induced by this multivalent lectin.

  16. Expression and modulation of nerve growth factor in murine keratinocytes (PAM 212)

    SciTech Connect

    Tron, V.A.; Coughlin, M.D.; Jang, D.E.; Stanisz, J.; Sauder, D.N. )

    1990-04-01

    Nerve growth factor (NGF) is a polypeptide that is required for normal development and maintenance of the sympathetic and sensory nervous systems. Skin has been shown to contain relatively high amounts of NGF, which is in keeping with the finding that the quantity of NGF in a tissue is proportional to the extent of sympathetic innervation of that organ. Since the keratinocyte, a major cellular constituent of the skin, is known to produce other growth factors and cytokines, our experiments were designed to determine whether keratinocytes are a source of NGF. Keratinocyte-conditioned media from the keratinocyte cell line PAM 212 contained NGF-like activity, approximately 2-3 ng/ml, as detected by the neurite outgrowth assay. Freshly isolated BALB/c keratinocytes contained approximately 0.1 ng/ml. Using a cDNA probe directed against NGF, we demonstrated the presence of a 1.3-kb NGF mRNA in both PAM 212 and BALB/c keratinocytes. Since ultraviolet radiation (UV) is a potentially important modulating factor for cytokines in skin, we examined the effect of UV on NGF mRNA expression. Although UV initially inhibited the expression of keratinocyte NGF mRNA (4 h), by 24 h an induction of NGF mRNA was seen. The NGF signal could also be induced by phorbol esters. Thus, keratinocytes synthesize and express NGF, and its expression is modulated by UVB and phorbol esters.

  17. Cobra Venom Factor and Ketoprofen Abolish the Antitumor Effect of Nerve Growth Factor from Cobra Venom.

    PubMed

    Osipov, Alexey V; Terpinskaya, Tatiana I; Kuznetsova, Tatiana E; Ryzhkovskaya, Elena L; Lukashevich, Vladimir S; Rudnichenko, Julia A; Ulashchyk, Vladimir S; Starkov, Vladislav G; Utkin, Yuri N

    2017-09-06

    We showed recently that nerve growth factor (NGF) from cobra venom inhibited the growth of Ehrlich ascites carcinoma (EAC) inoculated subcutaneously in mice. Here, we studied the influence of anti-complementary cobra venom factor (CVF) and the non-steroidal anti-inflammatory drug ketoprofen on the antitumor NGF effect, as well as on NGF-induced changes in EAC histological patterns, the activity of lactate and succinate dehydrogenases in tumor cells and the serum level of some cytokines. NGF, CVF and ketoprofen reduced the tumor volume by approximately 72%, 68% and 30%, respectively. The antitumor effect of NGF was accompanied by an increase in the lymphocytic infiltration of the tumor tissue, the level of interleukin 1β and tumor necrosis factor α in the serum, as well as the activity of lactate and succinate dehydrogenases in tumor cells. Simultaneous administration of NGF with either CVF or ketoprofen abolished the antitumor effect and reduced all other effects of NGF, whereas NGF itself significantly decreased the antitumor action of both CVF and ketoprofen. Thus, the antitumor effect of NGF critically depended on the status of the immune system and was abolished by the disturbance of the complement system; the disturbance of the inflammatory response canceled the antitumor effect as well.

  18. Shape changes induced by biologically active peptides and nerve growth factor in blood platelets of rabbits.

    PubMed

    Gudat, F; Laubscher, A; Otten, U; Pletscher, A

    1981-11-01

    1 Nerve growth factor (NGF), substance P (SP) and thymopoietin all caused shape change reactions of rapid onset in rabbit platelets. NGF had the highest maximal effect, and SP the lowest EC50 (concentration causing half maximal shape change). The action of SP was reversible within 5 min, whereas that of NGF lasted for at least 1 h. A series of other peptides were inactive. 2 After preincubation of platelets with SP, a second application of SP no longer caused a shape change reaction, whereas the effect of NGF was not influenced. 3 An oxidized NGF-derivative without biological activity did not cause a shape change reaction, neither did epidermal growth factor. 4 Prostaglandin E1 (PGE1) and pretreatment of the platelets with 3% butanol, which counteract the shape changes caused by 5-hydroxytryptamine (5-HT) and adenosine 3',5'-diphosphate, also antagonized those induced by NGF and SP. Neither heparin nor methysergide, an antagonist of 5-HT-receptors, influenced the shape change induced by NGF or SP. The action of NGF was also antagonized by a specific antibody to NGF. 5 Thymopoietin, like the basic polypeptide polyornithine (mol. wt. 40,000) was not antagonized by PGE1 and butanol. Heparin, which counteracted the effect of polyornithine, did not influence that of thymopoietin. 6 In conclusion, different modes of action are involved in the shape change of blood platelets induced by polypeptides and proteins. SP and NGF may act by stimulating specific membrane receptors.

  19. Nerve growth factor modulate proliferation of cultured rabbit corneal endothelial cells and epithelial cells.

    PubMed

    Li, Xinyu; Li, Zhongguo; Qiu, Liangxiu; Zhao, Changsong; Hu, Zhulin

    2005-01-01

    In order to investigate the effect of nerve growth factor (NGF) on the proliferation of rabbit corneal endothelial cells and epithelial cells, the in vitro cultured rabbit corneal endothelial cells and epithelial cells were treated with different concentrations of NGF. MTT assay was used to examine the clonal growth and proliferation of the cells by determining the absorbency values at 570 nm. The results showed that NGF with three concentrations ranging from 5 U/mL to 500 U/mL enhanced the proliferation of rabbit corneal endothelial cells in a concentration-dependent manner. 50 U/mL and 500 U/mL NGF got more increase of proliferation than that of 5 U/mL NGF did. Meanwhile, 50 U/mL and 500 U/mL NGF could promote the proliferation of the rabbit corneal epithelial cells significantly in a concentration-dependent manner. However, 5 U/mL NGF did not enhance the proliferation of epithelial cells. It was suggested that exogenous NGF can stimulate the proliferation of both rabbit corneal endothelial and epithelial cells, but the extent of modulation is different.

  20. Reduced maternal and cord nerve growth factor levels in preterm deliveries.

    PubMed

    Dhobale, Madhavi; Mehendale, Savita; Pisal, Hemlata; Nimbargi, Vandana; Joshi, Sadhana

    2012-04-01

    Nerve growth factor (NGF) is a neurotrophin, which exerts an important role in the development and function of the central and peripheral nervous system. There is limited information regarding the levels of NGF during pregnancy and its role in fetal development. We have earlier reported increased oxidative stress in pregnancy complications. The present study examines the levels of NGF in maternal and cord samples in preterm deliveries and its association with oxidative stress marker. A total number of 96 women delivering preterm (<37 weeks gestation) and 94 women delivering at term (control group) (≥37 weeks gestation) were recruited. Plasma NGF levels were measured in both mother and cord plasma using the Emax Immuno Assay System Promega kit. Maternal and cord plasma NGF levels were significantly reduced (p<0.05 for both) in women delivering preterm as compared to term. There was a positive association between maternal and cord plasma NGF levels (p=0.022). Maternal NGF levels were negatively (p=0.017) associated with maternal malondialdehyde (MDA) levels. Reduced cord NGF levels may affect fetal growth in preterm deliveries which may have implications for the neurodevelopmental pathologies in later life. Circulating maternal NGF levels in preterm pregnancies may be a useful marker to predict NGF levels in the neonate.

  1. Nerve Growth Factor Signaling from Membrane Microdomains to the Nucleus: Differential Regulation by Caveolins

    PubMed Central

    Spencer, Ambre; Yu, Lingli; Guili, Vincent; Reynaud, Florie; Ding, Yindi; Ma, Ji; Jullien, Jérôme; Koubi, David; Gauthier, Emmanuel; Cluet, David; Falk, Julien; Castellani, Valérie; Yuan, Chonggang; Rudkin, Brian B.

    2017-01-01

    Membrane microdomains or “lipid rafts” have emerged as essential functional modules of the cell, critical for the regulation of growth factor receptor-mediated responses. Herein we describe the dichotomy between caveolin-1 and caveolin-2, structural and regulatory components of microdomains, in modulating proliferation and differentiation. Caveolin-2 potentiates while caveolin-1 inhibits nerve growth factor (NGF) signaling and subsequent cell differentiation. Caveolin-2 does not appear to impair NGF receptor trafficking but elicits prolonged and stronger activation of MAPK (mitogen-activated protein kinase), Rsk2 (ribosomal protein S6 kinase 2), and CREB (cAMP response element binding protein). In contrast, caveolin-1 does not alter initiation of the NGF signaling pathway activation; rather, it acts, at least in part, by sequestering the cognate receptors, TrkA and p75NTR, at the plasma membrane, together with the phosphorylated form of the downstream effector Rsk2, which ultimately prevents CREB phosphorylation. The non-phosphorylatable caveolin-1 serine 80 mutant (S80V), no longer inhibits TrkA trafficking or subsequent CREB phosphorylation. MC192, a monoclonal antibody towards p75NTR that does not block NGF binding, prevents exit of both NGF receptors (TrkA and p75NTR) from lipid rafts. The results presented herein underline the role of caveolin and receptor signaling complex interplay in the context of neuronal development and tumorigenesis. PMID:28338624

  2. Mammalian Target of Rapamycin (mTOR) Activation Increases Axonal Growth Capacity of Injured Peripheral Nerves*

    PubMed Central

    Abe, Namiko; Borson, Steven H.; Gambello, Michael J.; Wang, Fan; Cavalli, Valeria

    2010-01-01

    Unlike neurons in the central nervous system (CNS), injured neurons in the peripheral nervous system (PNS) can regenerate their axons and reinnervate their targets. However, functional recovery in the PNS often remains suboptimal, especially in cases of severe damage. The lack of regenerative ability of CNS neurons has been linked to down-regulation of the mTOR (mammalian target of rapamycin) pathway. We report here that PNS dorsal root ganglial neurons (DRGs) activate mTOR following damage and that this activity enhances axonal growth capacity. Furthermore, genetic up-regulation of mTOR activity by deletion of tuberous sclerosis complex 2 (TSC2) in DRGs is sufficient to enhance axonal growth capacity in vitro and in vivo. We further show that mTOR activity is linked to the expression of GAP-43, a crucial component of axonal outgrowth. However, although TSC2 deletion in DRGs facilitates axonal regrowth, it leads to defects in target innervation. Thus, whereas manipulation of mTOR activity could provide new strategies to stimulate nerve regeneration in the PNS, fine control of mTOR activity is required for proper target innervation. PMID:20615870

  3. Synergistic induction of neurite outgrowth by nerve growth factor or epidermal growth factor and interleukin-6 in PC12 cells.

    PubMed

    Wu, Y Y; Bradshaw, R A

    1996-05-31

    Native PC12 cells respond differentially to nerve growth factor (NGF) but not interleukin-6 (IL-6); PC12-E2 cells, a stable variant, respond to both stimuli (and more rapidly to NGF). Neither responds to epidermal growth factor (EGF). NGF primarily induces the RAS/extracellular signal-regulated kinase (ERK) pathway and IL-6 activates a JAK (Janus tyrosine kinase)/STAT (signal transducers and activators of transcription) response. EGF also stimulates RAS/ERK but in a transient manner. When either cell type is treated with combinations of NGF, EGF, and IL-6, at concentrations that produce modest or no response, a substantial augmentation of neurite outgrowth is observed. With PC12-E2 cells, a subthreshold concentration of IL-6 increases NGF response by approximately 2-3-fold after 1-2 days; the increase with EGF is more pronounced. Native PC12 cells show even greater synergistic effects with NGF and IL-6. The most dramatic effect was observed with low levels of EGF, where IL-6 increased the percentage of responsive cells from zero to approximately 60% after 3 days. In addition, two neural-specific transcripts, GAP-43 and SCG-10, are synergistically increased by the combinations of growth factors. Importantly, IL-6 does not enhance ERK phosphorylation in the presence of either NGF or EGF. In contrast, NGF and EGF, in the presence or absence of IL-6, cause mobility shifts of Stat3 that are consistent with serine phosphorylations. Although these modifications do not lead to activation and translocation by themselves, in the presence of the tyrosine phosphorylation induced by IL-6, they may play a role in the synergistic responses. These observations suggest a differentially regulated two-stage mechanism for the differentiative response of PC12 cells to NGF.

  4. Membrane proteins of the nerve growth cone and their developmental regulation

    SciTech Connect

    Simkowitz, P.; Ellis, L.; Pfenninger, K.H.

    1989-03-01

    The membrane polypeptides of growth cone fragments (growth cone particles, GCPs) isolated from fetal rat brain by subcellular fractionation have been analyzed in further detail. The major polypeptides of salt-washed GCP membranes detected by 1-dimensional gel electrophoresis resolve in 2-dimensional gels as a spot of 52 kDa that comigrates with beta-tubulin and reacts with anti-beta-tubulin; a 46 kDa, pl 4.3, polypeptide (pp46) that has no equivalent in the soluble fraction and is identical to one of the GCP's major phosphoproteins and to GAP43; a spot of 42 kDa that comigrates with actin; and a species of 34 kDa (p34) without soluble equivalent. The prominent 38 kDa doublet identified in 1-dimensional gels is difficult to resolve in 2-dimensional gels. The major phosphoproteins pp80ac, pp46, and pp40, as well as p34 partition into the oil phase of Triton X-114 extracts, suggesting that they are integral membrane proteins, at least in our experimental conditions. The properties of pp46 reported here are in conflict with the highly hydrophilic amino acid sequence predicted for GAP43/B50/F1. Growth-cone and presynaptic membrane proteins are compared as follows. After eye injection of 35S-methionine, GCPs and synaptosomes are isolated from the target areas of optic nerve of fetal and adult rats, respectively. Polypeptides are separated by 1- and 2-dimensional gel electrophoresis and the radiolabeled species identified fluorographically. The comparison of labeled GCP and synaptosome polypeptides shows that all 5 major Coomassie blue-stained polypeptides of GCP membranes (52, 46, 42, 38, 34 kDa) are intensely labeled after eye injection. However, in synaptosomes, these polypeptides are weakly labeled if at all; instead, an intensely labeled polypeptide of 28 kDa, and several additional species not seen in GCPs, have appeared.

  5. DNA methyltransferase 3a and mitogen-activated protein kinase signaling regulate the expression of fibroblast growth factor-inducible 14 (Fn14) during denervation-induced skeletal muscle atrophy.

    PubMed

    Tajrishi, Marjan M; Shin, Jonghyun; Hetman, Michal; Kumar, Ashok

    2014-07-18

    The TWEAK-fibroblast growth factor-inducible 14 (Fn14) system is a critical regulator of denervation-induced skeletal muscle atrophy. Although the expression of Fn14 is a rate-limiting step in muscle atrophy on denervation, mechanisms regulating gene expression of Fn14 remain unknown. Methylation of CpG sites within promoter region is an important epigenetic mechanism for gene silencing. Our study demonstrates that Fn14 promoter contains a CpG island close to transcription start site. Fn14 promoter also contains multiple consensus DNA sequence for transcription factors activator protein 1 (AP1) and specificity protein 1 (SP1). Denervation diminishes overall genomic DNA methylation and causes hypomethylation at specific CpG sites in Fn14 promoter leading to the increased gene expression of Fn14 in skeletal muscle. Abundance of DNA methyltransferase 3a (Dnmt3a) and its interaction with Fn14 promoter are repressed in denervated skeletal muscle of mice. Overexpression of Dnmt3a inhibits the gene expression of Fn14 and attenuates skeletal muscle atrophy upon denervation. Denervation also causes the activation of ERK1/2, JNK1/2, and ERK5 MAPKs and AP1 and SP1, which stimulate the expression of Fn14 in skeletal muscle. Collectively, our study provides novel evidence that Dnmt3a and MAPK signaling regulate the levels of Fn14 in skeletal muscle on denervation. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. Nerve Growth Factor Gene Therapy: Activation of Neuronal Responses in Alzheimer Disease.

    PubMed

    Tuszynski, Mark H; Yang, Jennifer H; Barba, David; U, Hoi-Sang; Bakay, Roy A E; Pay, Mary M; Masliah, Eliezer; Conner, James M; Kobalka, Peter; Roy, Subhojit; Nagahara, Alan H

    2015-10-01

    Alzheimer disease (AD) is the most common neurodegenerative disorder and lacks effective disease-modifying therapies. In 2001, we initiated a clinical trial of nerve growth factor (NGF) gene therapy in AD, the first effort at gene delivery in an adult neurodegenerative disorder. This program aimed to determine whether a nervous system growth factor prevents or reduces cholinergic neuronal degeneration in patients with AD. We present postmortem findings in 10 patients with survival times ranging from 1 to 10 years after treatment. To determine whether degenerating neurons in AD retain an ability to respond to a nervous system growth factor delivered after disease onset. Patients in this anatomicopathological study were enrolled in clinical trials from March 2001 to October 2012 at the University of California, San Diego, Medical Center in La Jolla. Ten patients with early AD underwent NGF gene therapy using ex vivo or in vivo gene transfer. The brains of all 8 patients in the first phase 1 ex vivo trial and of 2 patients in a subsequent phase 1 in vivo trial were examined. Brains were immunolabeled to evaluate in vivo gene expression, cholinergic neuronal responses to NGF, and activation of NGF-related cell signaling. In 2 patients, NGF protein levels were measured by enzyme-linked immunosorbent assay. Among 10 patients, degenerating neurons in the AD brain responded to NGF. All patients exhibited a trophic response to NGF in the form of axonal sprouting toward the NGF source. Comparing treated and nontreated sides of the brain in 3 patients who underwent unilateral gene transfer, cholinergic neuronal hypertrophy occurred on the NGF-treated side (P < .05). Activation of cellular signaling and functional markers was present in 2 patients who underwent adeno-associated viral vectors (serotype 2)-mediated NGF gene transfer. Neurons exhibiting tau pathology and neurons free of tau expressed NGF, indicating that degenerating cells can be infected with therapeutic

  7. Effects of Nerve Growth Factor and Basic Fibroblast Growth Factor Promote Human Dental Pulp Stem Cells to Neural Differentiation.

    PubMed

    Zhang, Jinlong; Lian, Min; Cao, Peipei; Bao, Guofeng; Xu, Guanhua; Sun, Yuyu; Wang, Lingling; Chen, Jiajia; Wang, Yi; Feng, Guijuan; Cui, Zhiming

    2017-04-01

    Dental pulp stem cells (DPSCs) were the most widely used seed cells in the field of neural regeneration and bone tissue engineering, due to their easily isolation, lack of ethical controversy, low immunogenicity and low rates of transplantation rejection. The purpose of this study was to investigate the role of basic fibroblast growth factor (bFGF) and nerve growth factor (NGF) on neural differentiation of DPSCs in vitro. DPSCs were cultured in neural differentiation medium containing NGF and bFGF alone or combination for 7 days. Then neural genes and protein markers were analyzed using western blot and RT-PCR. Our study revealed that bFGF and NGF increased neural differentiation of DPSCs synergistically, compared with bFGF and NGF alone. The levels of Nestin, MAP-2, βIII-tubulin and GFAP were the most highest in the DPSCs + bFGF + NGF group. Our results suggested that bFGF and NGF signifiantly up-regulated the levels of Sirt1. After treatment with Sirt1 inhibitor, western blot, RT-PCR and immunofluorescence staining showed that neural genes and protein markers had markedly decreased. Additionally, the ERK and AKT signaling pathway played a key role in the neural differentiation of DPSCs stimulated with bFGF + NGF. These results suggested that manipulation of the ERK and AKT signaling pathway may be associated with the differentiation of bFGF and NGF treated DPSCs. Our date provided theoretical basis for DPSCs to treat neurological diseases and repair neuronal damage.

  8. Nerve growth factor (NGF)-conjugated electrospun nanostructures with topographical cues for neuronal differentiation of mesenchymal stem cells.

    PubMed

    Cho, Young Il; Choi, Ji Suk; Jeong, Seo Young; Yoo, Hyuk Sang

    2010-12-01

    Mesenchymal stem cells (MSCs) were cultivated on the surface of nerve growth factor (NGF)-conjugated aligned nanofibrous meshes for neuronal differentiation. Amine-terminated poly(ethylene glycol) was conjugated to poly(ε-caprolactone) to prepare amine-functionalized block copolymers. The synthesized polymer was electrospun in a rotating drum to prepare aligned nanofibrous meshes. A nerve growth factor was chemically immobilized on the surface-exposed amine groups of the electrospun nanofibrous meshes in the aqueous phase. In vitro release profiles of the nerve growth factor were investigated for NGF-immobilized nanofibrous meshes. The conjugated nerve growth factor was not released for 7 days, while the growth factor physically adsorbed on the nanofibrous meshes showed an initial burst release. MSCs were cultivated on the NGF-conjugated nanofibrous meshes for 5 days, and total RNA was extracted from the cultivated cells. mRNA was extracted from cells for measuring expression levels of neuronal differentiation markers, including nestin, tubulin βIII and map2, in the cultivated stem cells. The conjugation of NGF significantly increased the expression levels of the marker proteins for neuron cells while physically adsorbed NGFs on nanofibrous meshes showed low expression of these marker genes. Furthermore, alignments of nanofibrous meshes clearly increased the expression levels of neuronal makers while the nanofibrous mesh without the topographical cue did not affect neuronal differentiation of the cultivated stem cells. Confocal microscopy revealed that the stem cells on the NGF-conjugated aligned nanofibrous meshes showed intense staining with antibodies against neuronal makers as well as elongated morphology compared to other groups. Thus, the NGF-conjugated nanofibrous meshes with topographical cues significantly increased the neuronal differentiation of mesenchymal stem cells in comparison to NGF-adsorbed nanofibrous meshes.

  9. Neuroprotection by Cocktails of Dietary Antioxidants under Conditions of Nerve Growth Factor Deprivation.

    PubMed

    Amara, Flavio; Berbenni, Miluscia; Fragni, Martina; Leoni, Giampaolo; Viggiani, Sandra; Ippolito, Vita Maria; Larocca, Marilena; Rossano, Rocco; Alberghina, Lilia; Riccio, Paolo; Colangelo, Anna Maria

    2015-01-01

    Dietary antioxidants may be useful in counteracting the chronic inflammatory status in neurodegenerative diseases by reducing oxidative stress due to accumulation of reactive oxygen species (ROS). In this study, we newly described the efficacy of a number of dietary antioxidants (polyphenols, carotenoids, thiolic compounds, and oligoelements) on viability of neuronal PC12 cells following Nerve Growth Factor (NGF) deprivation, a model of age-related decrease of neurotrophic support that triggers neuronal loss. Neuroprotection by antioxidants during NGF deprivation for 24 h was largely dependent on their concentrations: all dietary antioxidants were able to efficiently support cell viability by reducing ROS levels and restoring mitochondrial function, while preserving the neuronal morphology. Moreover, ROS reduction and neuroprotection during NGF withdrawal were also achieved with defined cocktails of 3-6 different antioxidants at concentrations 5-60 times lower than those used in single treatments, suggesting that their antioxidant activity was preserved also at very low concentrations. Overall, these data indicate the beneficial effects of antioxidants against oxidative stress induced by decreased NGF availability and suggest that defined cocktails of dietary factors at low concentrations might be a suitable strategy to reduce oxidative damage in neurodegenerative diseases, while limiting possible side effects.

  10. Nerve growth factor-endothelial cell interaction leads to angiogenesis in vitro and in vivo.

    PubMed

    Cantarella, Giuseppina; Lempereur, Laurence; Presta, Marco; Ribatti, Domenico; Lombardo, Gabriella; Lazarovici, Philip; Zappalà, Giovanna; Pafumi, Carlo; Bernardini, Renato

    2002-08-01

    Nerve growth factor (NGF) has important functions during embryonic development and on various tissues and organs under normal and pathological conditions during the extrauterine life. RT-PCR analysis and immunological methods demonstrate that human umbilical vein endothelial cells (HUVECs) express the NGF receptors trkA(NGFR) and p75NTR. NGF treatment caused a rapid phosphorylation of trkA(NGFR) in HUVECs, determining a parallel increase of phosphorylated ERK1/2. Accordingly, NGF induced a significant increase in HUVEC proliferation that was abolished by the trkA(NGFR) inhibitor K252a. Also, HUVECs express significant levels of NGF under standard culture conditions that were up-regulated during serum starvation. Endogenous NGF was responsible for the basal levels of trkA(NGFR) and ERK1/2 phosphorylation observed in untreated HUVEC cultures. Finally, NGF exerted a potent, direct, angiogenic activity in vivo when delivered onto the chorioallantoic membrane of the chicken embryo. The data indicate that NGF may play an important role in blood vessel formation in the nervous system and in several pathological processes, including tumors and inflammatory diseases. Unraveling mechanisms of NGF-dependent angiogenesis could provide valuable tools for novel therapeutic approaches in antiangiogenic therapy.

  11. Neuromodulatory loop mediated by nerve growth factor and interleukin 6 in thymic stromal cell cultures.

    PubMed Central

    Screpanti, I; Meco, D; Scarpa, S; Morrone, S; Frati, L; Gulino, A; Modesti, A

    1992-01-01

    Neural crest cell derivatives have been suggested to be involved in thymus development. We established nonlymphoid thymic stromal cell cultures capable of supporting T-cell differentiation. In these nonlymphoid cell cultures, we identified cells with phenotypic and biochemical markers specific for neuronal cells. Neurofilament mRNA and 68- and 160-kDa neurofilament proteins, as well as 74-kDa synapsin I isoform, were expressed in many of the cultured cells. For example, neurofilament immunoreactivity was detected in 20-30% of the cells. To see whether thymic neuronal-like cells were involved in a neural differentiation pathway, we investigated the effect of nerve growth factor (NGF) and interleukin 6 (IL-6), two known neurotrophic factors. The expression of the above-described neural markers was enhanced by NGF and IL-6, which we report to be produced in an autocrine way by thymic stromal cell cultures. Finally, we found that IL-6 gene expression in these cell cultures was enhanced by NGF. Evidence is thus offered of a neuromodulatory loop within the thymic stromal cell population supported by local production of NGF and IL-6 and involving neural cell elements. Interestingly, IL-6, which is known to be implicated in thymocyte differentiation, also displays a neuromodulatory activity on thymic stromal cells, suggesting a multivalent role for this cytokine within the thymus. Images PMID:1373490

  12. Nerve growth factor serum concentrations rise after successful cognitive-behavioural therapy of generalized anxiety disorder.

    PubMed

    Jockers-Scherübl, Maria C; Zubraegel, Doris; Baer, Thomas; Linden, Michael; Danker-Hopfe, Heidi; Schulte-Herbrüggen, Olaf; Neu, Peter; Hellweg, Rainer

    2007-01-30

    Generalized anxiety disorder (GAD) is a chronic stress disease with permanent physical tension and cognitive strain. Raised nerve growth factor (NGF) serum levels were reported as an acute stress reaction in soldiers before their first parachute jump even before the rise in cortisol. Taking GAD as a clinical model of chronic stress, we measured NGF in the serum of 22 patients with GAD before and after cognitive-behavioural therapy (CBT) and compared them to those of healthy normal controls. Treatment response was tested by the values of the State and Trait of Anxiety Inventory (STAI) and the Hamilton Anxiety Scale (HAM-A) as treatment outcome variables. The NGF values of patients and controls were similar at baseline (p=0.8941); however, with successful treatment, corresponding to a mean reduction in the HAM-A by more than 50% and a reduction in the clinical global impression scale (CGI) median from 4 to 1, the patients' NGF serum concentrations rose significantly (p=0.0006) which might correspond to an altered stress reaction, possibly contributing to good therapeutic response with CBT. There were 3 patients with a HAM-A decrease of less than 15%. In those patients NGF rose only marginally. Hence, the increase in serum NGF seems to indicate good treatment response.

  13. Bilateral choroidal neovascularization associated with optic nerve head drusen treated by antivascular endothelial growth factor therapy

    PubMed Central

    Delas, Barbara; Almudí, Lorena; Carreras, Anabel; Asaad, Mouafk

    2012-01-01

    Objective To report a good clinical outcome in a patient with bilateral choroidal neovascularization (CNV) associated with optic nerve head drusen (ONHD) treated with intravitreal ranibizumab injection. Methods A 12-year-old girl was referred for loss of right eye vision detected in a routine check-up. Best-corrected visual acuity (BCVA) was hand movements in the right eye and 0.9 in the left eye. Funduscopy revealed the presence of superficial and buried bilateral ONHD, which was confirmed by ultrasonography and computed tomography, and the study was completed with perimetry. The presence of bilateral CNV, active in the right eye, was observed and subsequently confirmed using fluorescein angiography and optical coherence tomography. Results Treatment with two consecutive injections of intravitreal ranibizumab resulted in inactivation of the neovascular membrane with subretinal fluid reabsorption and improved right eye BCVA. After 12 months’ follow-up, this was 20/60 and stable. Conclusion Although there are no published studies of safety in children, antiangiogenic therapy for CNV secondary to ONHD may be useful and safe. A search of the literature produced only one previously reported case of ONHD-associated CNV treated with antivascular endothelial growth factor alone. PMID:22368440

  14. Nerve growth factor metabolic dysfunction in Alzheimer's disease and Down syndrome.

    PubMed

    Iulita, M Florencia; Cuello, A Claudio

    2014-07-01

    Alzheimer's disease (AD) is a devastating neurodegenerative condition and the most common type of amnestic dementia in the elderly. Individuals with Down syndrome (DS) are at increased risk of developing AD in adulthood as a result of chromosome 21 trisomy and triplication of the amyloid precursor protein (APP) gene. In both conditions, the central nervous system (CNS) basal forebrain cholinergic system progressively degenerates, and such changes contribute to the manifestation of cognitive decline and dementia. Given the strong dependency of these neurons on nerve growth factor (NGF), it was hypothesized that their atrophy was caused by NGF deficits. However, in AD, the synthesis of NGF is not affected at the transcript level and there is a marked increase in its precursor, proNGF. This apparent paradox remained elusive for many years. In this review, we discuss the recent evidence supporting a CNS deficit in the extracellular metabolism of NGF, both in AD and in DS brains. We describe the nature of this trophic disconnection and its implication for the atrophy of basal forebrain cholinergic neurons. We further discuss the potential of NGF pathway markers as diagnostic indicators of a CNS trophic disconnection.

  15. Serum levels of nerve growth factor (NGF) in patients with major depression disorder and suicide risk.

    PubMed

    Wiener, Carolina David; de Mello Ferreira, Sharon; Pedrotti Moreira, Fernanda; Bittencourt, Guilherme; de Oliveira, Jacqueline Flores; Lopez Molina, Mariane; Jansen, Karen; de Mattos Souza, Luciano Dias; Rizzato Lara, Diogo; Portela, Luiz Valmor; da Silva, Ricardo Azevedo; Oses, Jean Pierre

    2015-09-15

    Nerve growth factor (NGF) is an important member of the neurotrophins group and their involvement in the pathophysiology of major depression disorder (MDD) and suicide risk (SR) has been recently suggested. The aim of this study is to evaluate the changes in NGF serum levels in individuals with MDD and with or without risk of suicide, in subjects from a young population-based sample. This is a paired cross-sectional study nested in a population-based study. Individuals were rated for MDD and SR by a diagnostic interview--Mini International Neuropsychiatric Interview (M.I.N.I). The total population of the sample was comprised of 141 subjects distributed in three groups: 47 healthy controls, 47 subjects with current depressive episode without SR (MDD) and 47 subjects with current depressive episode and with SR (MDD + SR). NGF serum levels were significantly reduced in the MDD and MDD + SR groups when compared with controls (p ≤ 0.001). However, there were no differences in NGF levels between the MDD and MDD + SR groups (p = 1.000). These results suggest that reduced NGF serum levels can be a possible biomarker of MDD.

  16. Nerve growth factor modulates TRPV1 expression and function and mediates pain in chronic pancreatitis.

    PubMed

    Zhu, Yaohui; Colak, Tugba; Shenoy, Mohan; Liu, Liansheng; Pai, Reetesh; Li, Cuiping; Mehta, Kshama; Pasricha, Pankaj Jay

    2011-07-01

    The pathogenesis of pain in chronic pancreatitis (CP) is poorly understood and treatment remains difficult. We hypothesized that nerve growth factor (NGF) plays a key role in this process via its effects on the transient receptor potential vanilloid 1, TRPV1. CP was induced by intraductal injection of trinitrobenzene sulfonic acid in rats. After 3 weeks, anti-NGF antibody or control serum was administered daily for 1 week. Pancreatic hyperalgesia was assessed by nocifensive behavioral response to electrical stimulation of the pancreas as well as by referred somatic pain assessed by von Frey filament testing. TRPV1 currents in pancreatic sensory neurons were examined by patch-clamp. The expression and function of TRPV1 in pancreas-specific nociceptors was examined by immunostaining and quantification of messenger RNA levels. Blockade of NGF significantly attenuated pancreatic hyperalgesia and referred somatic pain compared with controls. It also decreased TRPV1 current density and open probability and reduced the proportion of pancreatic sensory neurons that expressed TRPV1 as well as levels of TRPV1 in these neurons. These findings emphasize a key role for NGF in pancreatic pain and highlight the role it plays in the modulation of TRPV1 expression and activity in CP. Copyright © 2011 AGA Institute. Published by Elsevier Inc. All rights reserved.

  17. Nerve growth factor preserves a critical motor period in rat striatum.

    PubMed

    Wolansky, M J; Paratcha, G C; Ibarra, G R; Azcurra, J M

    1999-01-01

    We previously found the occurrence of a critical motor period during rat postnatal development where circling training starting the 7-day schedule at 30 days-but not before or after-induces a lifetime drop in the binding to cholinergic muscarinic receptors (mAChRs) in striatum. Here, we studied whether nerve growth factor (NGF) participates in this restricted period of muscarinic sensitivity. For this purpose, we administered mouse salival gland 2.5S NGF (1.4 or 0.4 microg/day, infused by means of ALZA minipumps) by intrastriatal unilateral route between days 25 and 39, and then trained rats starting at 40 days. Under these conditions, NGF induced a long-term reduction in the striatal [3H] quinuclidilbenzylate (QNB) binding sites despite the fact that motor training was carried out beyond the natural critical period. Thus, at day 70, measurement of specific QNB binding in infused striata of trained rats showed decreases of 42% (p < .0004) and 33% (p < .02) after administration of the higher and lower NGF doses, respectively, with respect to trained rats treated with cytochrome C, for control. Noncannulated striata of the NGF-treated rats also showed a decrease in QNB binding sites (44%; p < .0001) only at the higher infusion rate. This effect was not found in the respective control groups. Our observations show that NGF modulates the critical period in which activity-dependent mAChR setting takes place during rat striatal maturation.

  18. Developmentally Regulated Expression of the Nerve Growth Factor Receptor Gene in the Periphery and Brain

    NASA Astrophysics Data System (ADS)

    Buck, C. R.; Martinez, Humberto J.; Black, Ira B.; Chao, Moses V.

    1987-05-01

    Nerve growth factor (NGF) regulates development and maintenance of function of peripheral sympathetic and sensory neurons. A potential role for the trophic factor in brain has been detected only recently. The ability of a cell to respond to NGF is due, in part, to expression of specific receptors on the cell surface. To study tissue-specific expression of the NGF receptor gene, we have used sensitive cRNA probes for detection of NGF receptor mRNA. Our studies indicate that the receptor gene is selectively and specifically expressed in sympathetic (superior cervical) and sensory (dorsal root) ganglia in the periphery, and by the septum-basal forebrain centrally, in the neonatal rat in vivo. Moreover, examination of tissues from neonatal and adult rats reveals a marked reduction in steady-state NGF receptor mRNA levels in sensory ganglia. In contrast, a 2- to 4-fold increase was observed in the basal forebrain and in the sympathetic ganglia over the same time period. Our observations suggest that NGF receptor mRNA expression is developmentally regulated in specific areas of the nervous system in a differential fashion.

  19. The Effect of Vitamin D Treatment On Nerve Growth Factor (NGF) Release From Hippocampal Neurons

    PubMed Central

    GEZEN-AK, Duygu; DURSUN, Erdinç; YILMAZER, Selma

    2014-01-01

    Introduction Vitamin D, the main function of which is thought to be the maintenance of calcium and phosphate homeostasis and bone structure, has been shown in recent studies to have important roles in brain development as well. A certain vitamin D receptor (VDR) gene haplotype was reported, for the first time by our group, to increase the risk of developing Alzheimer’s disease. Our studies also showed that vitamin D prevents beta amyloid-induced calcium elevation and toxicity that target nerve growth factor (NGF) release in cortical neurons; beta amyloid suppresses VDR expression and the disruption of vitamin D-VDR pathway mimics beta amyloid-induced neurodegeneration. In this study, our aim was to investigate the effects of vitamin D on the NGF release from hippocampal neurons. Method Primary hippocampal neuron cultures that were prepared from 18-day-old Sprague-Dawley rat embryos were treated with vitamin D for 48 hours. The alteration in the NGF release was determined with ELISA. Cytotoxicity tests were also performed for all groups. Results The NGF release in vitamin D-treated group was significantly higher than in untreated control group. The protective effect of vitamin D against cytotoxicity was also observed. Conclusion Our results indicated that vitamin D regulates the release of NGF, a very important molecule for neuronal survival of hippocampal neurons as well as cortical neurons.

  20. Urinary nerve growth factor and a variable solifenacin dosage in patients with an overactive bladder.

    PubMed

    Ciftci, Seyfettin; Ozkurkcugil, Cuneyd; Yilmaz, Hasan; Ustuner, Murat; Yavuz, Ufuk; Yuksekkaya, Mustafa; Cekmen, Mustafa Baki

    2016-02-01

    We evaluated changes in urinary nerve growth factor (NGF) and NGF/creatinine (NGF/Cr) levels after increasing the dosage of solifenacin in overactive bladder patients. The study groups included 59 overactive bladder (OAB) patients and 20 healthy subjects as controls. We measured NGF at baseline for the patients and controls, and used the Overactive Bladder Awareness Tool (OAB-V8) to evaluate urinary symptoms. All patients received a treatment of solifenacin 5 mg for 6 weeks. The responders to treatment served as group 1 and nonresponders received solifenacin 10 mg for an additional 6 weeks. Responders and nonresponders to the 10-mg treatment were defined as groups 2 and 3 respectively. NGF was measured after each treatment using the ELISA method and normalized by the urinary creatinine levels (NGF/Cr). There were 21, 22 and 16 patients in groups 1, 2, and 3 respectively. At baseline, the NGF and NGF/Cr levels were higher in groups 1, 2, and 3 compared with the controls. After the solifenacin 5 mg treatment, the NGF and NGF/Cr levels of group 1 individuals decreased to those of the control level. After increasing the dosage of solifenacin to 10 mg in group 2, the NGF and NGF/Cr levels decreased to normal levels. In group 3 (patients who did not responded to any treatment), these levels remained unchanged. Our results suggest that urinary NGF could be a potential biomarker for monitoring the treatment of symptoms in OAB patients who are treated with solifenacin.

  1. Expression of nerve growth factor and its receptor, tyrosine kinase receptor A, in rooster testes.

    PubMed

    Ma, Wei; Wang, Chunqiang; Su, Yuhong; Tian, Yumin; Zhu, Hongyan

    2015-10-01

    Nerve growth factor (NGF), which is required for the survival and differentiation of the nervous system, is also thought to play an important role in the development of mammalian reproductive tissues. To explore the function of NGF in the male reproductive system of non-mammalian animals, we determined the presence of NGF and its receptor, tyrosine kinase receptor A (TrkA), in rooster testes and investigated the regulation of NGF and TrkA expression by follicle-stimulating hormone (FSH). The mRNA and protein levels of NGF and TrkA in 6-week-old rooster testes were lower than those in 12-, 16- or 20-week age groups; levels were highest in the 16-week group. Immunohistochemistry showed that NGF and TrkA were both detected in spermatogonia, spermatocytes and spermatids. NGF immunoreactivity was observed in Leydig cells and strong TrkA signals were present in Sertoli cells. Meanwhile, FSH increased TrkA transcript levels in rooster testes in a dose-dependent manner. We present novel evidence for the developmental and FSH-regulated expression of the NGF/TrkA system, and our findings suggest that the NGF/TrkA system may play a prominent role in chicken spermatogenesis. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Nerve growth factor evokes hyperalgesia in mice lacking the low-affinity neurotrophin receptor p75.

    PubMed

    Bergmann, I; Reiter, R; Toyka, K V; Koltzenburg, M

    1998-10-16

    Endogenous nerve growth factor (NGF) has been shown to be an important mediator of inflammatory pain and exogenous application of recombinant human NGF (rhNGF) produces pain and hyperalgesia in animals and humans. Since NGF can act through two receptors types, the high affinity tyrosine kinase A (trkA) receptor and the low affinity p75 receptor, we used transgenic mice lacking p75 to analyse the relative importance of these receptors. After systemic injection of rhNGF (5 mg/ kg), pharmacokinetic studies revealed similar serum levels and elimination profiles of exogenously administered rhNGF in both strains of mice. Although animals lacking p75 have increased mechanical and thermal withdrawal thresholds they developed both heat and mechanical hyperalgesia after systemic injection of rhNGF whose magnitude did not differ significantly from wildtype animals. This means that NGF-induced hyperalgesia can occur in the absence of the p75 receptor and suggests that the trkA receptor is sufficient to mediate the acute noxious action of NGF.

  3. Effects of Myoga on Memory and Synaptic Plasticity by Regulating Nerve Growth Factor-Mediated Signaling.

    PubMed

    Kim, Hyo Geun; Lim, Soonmin; Hong, Jongki; Kim, Ae-Jung; Oh, Myung Sook

    2016-02-01

    The flower bud of Zingiber mioga Roscoe, known as 'myoga' or Japanese ginger, has a pungent aroma and is commonly consumed as a spice, with pickles, or as a health supplement in Eastern Asia. Here, we evaluated the activity of myoga in the brain, focusing especially on nerve growth factor (NGF), which is believed to mediate synaptic plasticity, supporting learning and memory. In a rat primary hippocampal astrocyte culture system, treatment with myoga extract for 24 h significantly stimulated the production of NGF. In mice administered myoga extract for 14 days, 200 and 400 mg/kg/day treatment resulted in increased NGF levels in the hippocampus. Myoga extract treatment also regulated the phosphorylation of extracellular signal-regulated kinases and cAMP response element-binding protein in the mouse hippocampus, leading to increased synaptic plasticity. In addition, it significantly increased novel object recognition time and spontaneous alternation, indicating improvement in learning and memory. These results suggest that myoga helps regulate NGF and synaptic plasticity, increasing memory ability.

  4. Some characteristics of histamine secretion from rat peritoneal mast cells stimulated with nerve growth factor.

    PubMed Central

    Pearce, F L; Thompson, H L

    1986-01-01

    Nerve growth factor (NGF) isolated from mouse submandibular gland or from snake venom produced a dose-dependent release of histamine from isolated rat peritoneal mast cells. The response was almost totally dependent on the presence of extracellular calcium ions and on added phosphatidylserine or its lyso-derivative. At high concentrations, strontium ions could substitute for calcium. The process was non-cytotoxic, relatively slow, pH dependent and blocked by polyclonal antibodies to NGF. Binding of NGF to the mast cell was not dependent on added calcium. The release was unaffected by low molecular weight glucose polymers or specific quaternary ammonium salts and thus differed from that evoked by clinical dextran or polyamines. The release was not inhibited by soluble rat IgE or IgG and was unimpaired in mast cells recovered from specific pathogen free rats. As such it did not appear to be mediated through interaction with cell-fixed antibodies. The process further differed from anaphylactic histamine release in that there was no accompanying change in the intracellular level of adenosine 3',5'-cyclic monophosphate (cyclic AMP), the activated state induced by NGF was much more persistent than that evoked by antigen, and there was no cross-desensitization between the two latter stimuli. In total, these data suggest that NGF may induce secretion from rat mast cells by interaction with a specific receptor on the plasma membrane, possibly similar to that present on sensory and sympathetic neurones. PMID:2425086

  5. Serum nerve growth factor (NGF) levels in children with attention deficit/hyperactivity disorder (ADHD).

    PubMed

    Guney, Esra; Ceylan, Mehmet Fatih; Kara, Mehmet; Tekin, Neslihan; Goker, Zeynep; Senses Dinc, Gulser; Ozturk, Onder; Eker, Sevda; Kizilgun, Murat

    2014-02-07

    Attention deficit/hyperactivity disorder (ADHD) is the most commonly diagnosed neurobehavioral disorder of childhood. The etiopathogeny of ADHD has not been totally defined. Recent reports have suggested a pathophysiological role of neurotrophins in ADHD. In this study, we evaluated serum levels of nerve growth factor (NGF) in patients with ADHD. The sample population consisted of 44 child or adolescent patients diagnosed with ADHD according to DSM-IV criteria; 36 healthy subjects were included in the study as controls. Venous blood samples were collected, and NGF levels were measured. The mean serum NGF levels of the ADHD patients were significantly higher than those of the controls. Age and gender of the patients were not correlated with serum NGF levels. There were no significant differences in NGF levels among the combined and predominantly inattentive subtypes of ADHD. Our study suggests that there are higher levels of serum NGF in drug naive ADHD patients, and that increased levels of NGF might have an important role in the pathophysiology of ADHD. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  6. Depletion of nerve growth factor in chemotherapy-induced peripheral neuropathy associated with hematologic malignancies.

    PubMed

    Youk, Jeonghwan; Kim, Young-Sook; Lim, Jung-Ah; Shin, Dong-Yeop; Koh, Youngil; Lee, Soon-Tae; Kim, Inho

    2017-01-01

    To investigate whether the depletion of nerve growth factor (NGF) is associated with the development of chemotherapy-induced peripheral neuropathy (CIPN) in patients with hematologic malignancy. We prospectively enrolled hematologic cancer patients who had a plan to receive bortezomib, thalidomide, or vincristine. Baseline NGF levels were measured within one week before the start date of chemotherapy. Follow-up NGF levels were measured after four months from the start date of chemotherapy or the date when CIPN was initially diagnosed. Baseline and follow-up NGF pairs were measured in 45 patients (male/female = 27/18, median age = 63 years old). CIPN has developed in 28 patients. In the CIPN group, the level of NGF was significantly decreased after chemotherapy compared to the baseline (△NGF = -3.52 ±5.72; p-value = 0.003), while the NGF level of the no-CIPN group was not changed after chemotherapy. The differences in △NGF levels between the CIPN and no-CIPN group were more profound when analyzed in the subgroup of newly diagnosed multiple myeloma patients (△NGF = -4.14 ± 4.87 pg/ml for the CIPN group and +2.52 ± 8.39 pg/ml for the no-CIPN group; p-value = 0.043). This study shows that the depletion of NGF occurs during the development of CIPN, suggesting pathogenesis based on the role of NGF and therapeutic implications.

  7. Quantitative analysis of nerve growth factor in the amniotic fluid during chick embryonic development.

    PubMed

    Mashayekhi, Farhad; Dianati, Elham; Moghadam, Lotfali Masomi

    2011-04-01

    Nerve growth factor (NGF) and most neurotrophic factors support the proliferation and survival of particular types of neurons. Besidesthe pivotal role of NGF in the development of neuronal cells, it also has important functions on non-neuronal cells. The amnion surrounds the embryo, providing an aqueous environment for the embryo. A wide range of proteins has been identified in human amniotic fluid (AF). In this study, total protein concentration (TPC) and NGF level in AF samples from chick embryos were measured using a Bio-Rad protein assay, enzyme linked immunosorbent assay (ELISA) and Western blot. TPC increased from days E10 to day E18. There was a rapid increase in AF TPC on day E15 when compared to day E16. No significant changes in NGF levels have been seen from day E10 to day E14. There was a rapid increase in NGF content on days E15 and E16, and thereafter the levels decreased from day E16 to day E18. Since, NGF is important in brain development and changes in AF NGF levels have been seen in some CNS malformations, changes in the TPC and NGF levels in AF during chick embryonic development may be correlated with cerebral cortical development. It is also concluded that NGF is a constant component of the AF during chick embryogenesis.

  8. Differential activation of dendritic cells by nerve growth factor and brain-derived neurotrophic factor.

    PubMed

    Noga, O; Peiser, M; Altenähr, M; Knieling, H; Wanner, R; Hanf, G; Grosse, R; Suttorp, N

    2007-11-01

    Neurotrophins are involved in inflammatory reactions influencing several cells in health and disease including allergy and asthma. Dendritic cells (DCs) play a major role in the induction of inflammatory processes with an increasing role in allergic diseases as well. The aim of this study was to investigate the influence of neurotrophins on DC function. Monocyte-derived dendritic cells were generated from allergic and non-allergic donors. Neurotrophin receptors were demonstrated by western blotting, flow cytometry and fluorescence microscopy. Activation of small GTPases was evaluated by pull-down assays. DCs were incubated with nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) and supernatants were collected for measurement of IL-4, IL-6, IL-10, IL-12p70, TNF-alpha and TGF-beta. Receptor proteins were detectable by western blot, fluorescence activated cell sorting analysis and fluorescence microscopy. Signalling after neurotrophin stimulation occurred in a ligand-specific pattern. NGF led to decreased RhoA and increased Rac activation, while BDNF affected RhoA and Rac activity in a reciprocal fashion. Cells of allergics released a significantly increased amount of IL-6, while for healthy subjects a significantly higher amount of IL-10 was found. These data indicate that DCs are activated by the neurotrophins NGF and BDNF by different pathways in a receptor-dependant manner. These cells then may initiate inflammatory responses based on allergic sensitization releasing preferred cytokines inducing tolerance or a T-helper type 2 response.

  9. Neuroprotection by Cocktails of Dietary Antioxidants under Conditions of Nerve Growth Factor Deprivation

    PubMed Central

    Amara, Flavio; Berbenni, Miluscia; Fragni, Martina; Leoni, Giampaolo; Viggiani, Sandra; Ippolito, Vita Maria; Larocca, Marilena; Rossano, Rocco; Alberghina, Lilia; Riccio, Paolo; Colangelo, Anna Maria

    2015-01-01

    Dietary antioxidants may be useful in counteracting the chronic inflammatory status in neurodegenerative diseases by reducing oxidative stress due to accumulation of reactive oxygen species (ROS). In this study, we newly described the efficacy of a number of dietary antioxidants (polyphenols, carotenoids, thiolic compounds, and oligoelements) on viability of neuronal PC12 cells following Nerve Growth Factor (NGF) deprivation, a model of age-related decrease of neurotrophic support that triggers neuronal loss. Neuroprotection by antioxidants during NGF deprivation for 24 h was largely dependent on their concentrations: all dietary antioxidants were able to efficiently support cell viability by reducing ROS levels and restoring mitochondrial function, while preserving the neuronal morphology. Moreover, ROS reduction and neuroprotection during NGF withdrawal were also achieved with defined cocktails of 3–6 different antioxidants at concentrations 5–60 times lower than those used in single treatments, suggesting that their antioxidant activity was preserved also at very low concentrations. Overall, these data indicate the beneficial effects of antioxidants against oxidative stress induced by decreased NGF availability and suggest that defined cocktails of dietary factors at low concentrations might be a suitable strategy to reduce oxidative damage in neurodegenerative diseases, while limiting possible side effects. PMID:26236423

  10. Tiam1 as a Signaling Mediator of Nerve Growth Factor-Dependent Neurite Outgrowth

    PubMed Central

    Vilar, Marçal; Paratcha, Gustavo; Ledda, Fernanda

    2010-01-01

    Nerve Growth Factor (NGF)-induced neuronal differentiation requires the activation of members of the Rho family of small GTPases. However, the molecular mechanisms through which NGF regulates cytoskeletal changes and neurite outgrowth are not totally understood. In this work, we identify the Rac1-specific guanine exchange factor (GEF) Tiam1 as a novel mediator of NGF/TrkA-dependent neurite elongation. In particular, we report that knockdown of Tiam1 causes a significant reduction in Rac1 activity and neurite outgrowth induced by NGF. Physical interaction between Tiam1 and active Ras (Ras-GTP), but not tyrosine phosphorylation of Tiam1, plays a central role in Rac1 activation by NGF. In addition, our findings indicate that Ras is required to associate Tiam1 with Rac1 and promote Rac1 activation upon NGF stimulation. Taken together, these findings define a novel molecular mechanism through which Tiam1 mediates TrkA signaling and neurite outgrowth induced by NGF. PMID:20333299

  11. Zhangfei, a novel regulator of the human nerve growth factor receptor, trkA.

    PubMed

    Valderrama, Ximena; Rapin, Noreen; Misra, Vikram

    2008-10-01

    The replication of herpes simplex virus (HSV) in epithelial cells, and during reactivation from latency in sensory neurons, depends on a ubiquitous cellular protein called host cell factor (HCF). The HSV transactivator, VP16, which initiates the viral replicative cycle, binds HCF as do some other cellular proteins. Of these, the neuronal transcription factor Zhangfei suppresses the ability of VP16 to initiate the replicative cycle. It also suppresses Luman, another cellular transcription factor that binds HCF. Interactions of nerve growth factor (NGF) and its receptor tropomyosin-related kinase (trkA) appear to be critical for maintaining HSV latency. Because the neuronal transcription factor Brn3a, which regulates trkA expression, has a motif for binding HCF, we investigated if Zhangfei had an effect on its activity. We found that Brn3a required HCF for activating the trkA promoter and Zhangfei suppressed its activity in non-neuronal cells. However, in neuron-like NGF-differentiated PC12 cells, both Brn3a and Zhangfei activated the trkA promoter and induced the expression of endogenous trkA. In addition, capsaicin, a stressor, which activates HSV in in vitro models of latency, decreased levels of Zhangfei and trkA transcripts in NGF-differentiated PC12 cells.

  12. Altered expression of nerve growth factor and its receptors in the kidneys of diabetic rats.

    PubMed

    Aloe, Luigi; Rossi, Simona; Manni, Luigi

    2011-01-01

    Type 1 diabetes (DB) is a multifactorial metabolic disorder characterized by loss of insulin-producing pancreatic ß-cells, and metabolic and functional deficits in a number of cells, including kidney cells. Nerve growth factor (NGF) is a signaling molecule that is up-regulated in cells affected by diabetes-linked disorders. However, whether DB alters the expression of NGF in the kidney is not known. DB was induced in adult male rats with a single injection of streptozotocin (STZ), and NGF protein levels were analyzed in a time-course study in serum and kidney. The expression of NGF receptors in the kidneys of healthy and DB rats was evaluated by immunohistochemistry. NGF levels as well as apoptotic features in the kidneys of healthy rats injected with purified NGF were also assessed. This study revealed that DB elevates NGF levels in serum and NGF expression in the kidney and that subcutaneous administration of NGF causes a marked uptake of NGF in kidney cells. The elevated presence of NGF in kidney cells is not associated with proapoptotic factor expression. The present data suggest that NGF presence in the kidney might play a survival, and most probably protective, role in kidney cells.

  13. Nerve growth factor localization in the nasal mucosa of patients with persistent allergic rhinitis.

    PubMed

    Bresciani, M; Lalibertè, F; Lalibertè, M F; Gramiccioni, C; Bonini, S

    2009-01-01

    Nerve growth factor (NGF) and NGF receptors have been shown to be expressed by structural and infiltrating inflammatory cells in the human allergic bronchial mucosa and conjunctiva. In the nose, a positive immunostaining for NGF was recently reported in biopsies of subjects undergoing surgery for refractory nasal obstruction. This study was aimed at studying by immunohistochemistry NGF expression and localization in the nasal mucosa from subjects with moderate/severe persistent allergic rhinitis and natural allergen exposure. Immunostaining for NGF, tryptase and eosinophil cationic protein was performed in human nasal turbinate sections of 25 patients affected by persistent allergic rhinitis and sensitization to Dermatophagoides pteronyssinus. NGF was consistently expressed in the epithelium and in the submucosa of allergic rhinitic subjects, preferentially localized in eosinophils and mast cells. A strong NGF immunostaining was found in mucous cells of the epithelial lining and in the submucosal glands. As previously shown for allergic asthma and allergic conjunctivitis, NGF is also detectable in the nasal mucosa of patients with persistent allergic rhinitis. The preferential NGF localization in mucous cells of the epithelial lining and in the submucosal glands suggests a possible role for NGF in modulating secretion in allergic rhinitis and possibly other allergic diseases.

  14. The Multiple Life of Nerve Growth Factor: Tribute to Rita Levi-Montalcini (1909–2012)

    PubMed Central

    Aloe, Luigi; Chaldakov, George N.

    2013-01-01

    At the end of the 19th century, it was envisaged by Santiago Ramon y Cajal, but not, proven, that life at the neuronal level requires trophic support. The proof was obtained in the early 1950’s by work initiated by Rita Levi-Montalcini (RLM) discovering the nerve growth factor (NGF). Today, NGF and its relatives, collectively designated neurotrophins, are well recognized as mediators of multiple biological phenomena in health and disease, ranging from the neurotrophic through immunotrophic and epitheliotrophic to metabotrophic effects. Consequently, NGF and other neurotrophins are implicated in the pathogenesis of a large spectrum of neuronal and non-neuronal diseases, from Alzheimer’s and other neurodegenerative diseases to atherosclerosis and other cardiometabolic diseases. Recent studies demonstrated the therapeutic potentials of NGF in these diseases, including ocular and cutaneous diseases. Furthermore, NGF TrkA receptor antagonists emerged as novel drugs for pain, prostate and breast cancer, melanoma, and urinary bladder syndromes. Altogether, NGF’s multiple potential in health and disease is briefly described here. PMID:25207059

  15. Expression Profile of Nerve Growth Factor after Muscle Incision in the Rat

    PubMed Central

    Wu, Chaoran; Erickson, Mark A.; Xu, Jun; Wild, Kenneth D.; Brennan, Timothy J.

    2009-01-01

    Background Previous studies have demonstrated that nerve growth factor (NGF) is an important mediator of pathologic pain. Many studies have focused on cutaneous mechanisms for NGF-induced hyperalgesia; few have examined its contribution in deeper tissues like muscle. This study examined pain behaviors and the expression of NGF in incised hind paw flexor digitorum brevis muscle. Methods Adult Sprague-Dawley rats were pretreated with anti-NGF peptibody and underwent skin or skin plus deep fascia and muscle incision. Guarding pain behaviors were measured. Muscle NGF messenger RNA (mRNA) was measured by real time polymerase chain reaction. Changes in NGF protein expression were measured using western blot, enzyme-linked immunoabsorbent assay and immunohistochemistry. In situ hybridization for NGF mRNA was also performed. Results Pretreatment with anti-NGF peptibody (100 mg/kg) decreased the guarding behavior caused by deep fascia and muscle incision. Muscle NGF mRNA increased abruptly 2 h after incision and was the same as control by postoperative day 1. NGF protein increased from 4 h after incision, and was sustained for several days. NGF was localized in many calcitonin gene related peptide positive axons, few N52 positive axons, but not isolectin B4 positive axons in incised muscle. The sources of NGF mRNA included keratinocytes in epidermis and fibroblasts in deeper tissues. Conclusion Fibroblasts adjacent to the injury are sources of NGF in incised muscle. NGF is upregulated by incision of muscle and contributes to guarding pain behavior. PMID:19104181

  16. Effect of nerve growth factor (NGF) on the development of preimplantation rabbit embryos in vitro.

    PubMed

    Pei, Yijin

    2010-01-01

    This study aimed to investigate the effect of nerve growth factor (NGF) on the development of preimplantation rabbit embryos in vitro. Zygotes were collected from superovulated New Zealand rabbits 19 h after injection of hCG and immediately mating and cultured in TCM-199 plus fatty-acid free BSA with different concentrations of NGF. Zygotes not treated with NGF served as control. At 24 h, 48 h, 72 h and 96 h of the culture, the numbers of the early cleavage stage, morulae, blastocysts and hatching blastocysts were determined. The intrazonal diameter of the blastocyst and the total cell numbers per blastocyst were measured after 96 h of culture. The results showed: (1) NGF at 100 ng/mL and 1000 ng/mL could improve the numbers of the hatching blastocysts which developed compared to the control treatment (p < 0.05); (2) All concentrations of NGF increased the total cell numbers in the blastocysts compared to the control treatment (p < 0.05); (3) NGF had no significant effect on the blastocyst intrazonal diameter of the blastocysts at 96 h of culture (p = 0.493); (4) The proportion in the early cleavage stage at 24 h of culture (p = 0.635), of morulae at 48 h of culture (p = 0.812) and of blastocysts at 72 h of culture (p = 0.812) in all treatments were not significantly different.

  17. Memory-enhancing effect of Mori Fructus via induction of nerve growth factor.

    PubMed

    Kim, Hyo Geun; Oh, Myung Sook

    2013-07-14

    Fruits rich in phytochemicals have been shown to improve memory by protecting or enhancing neuronal functions mediated by neurotrophic factors, such as nerve growth factor (NGF), in the hippocampus. Mori Fructus (Morus alba L., Moraceae), also called mulberry, is used as a food, dietary supplement and an anti-ageing agent in traditional Oriental medicine. It is also known to contain abundant flavonoid compounds and to exhibit various pharmacological effects. The present study was performed to evaluate the memory-enhancing effect of Mori Fructus extract (ME) in mice, with a focus on NGF regulation. ME (20, 100 and 500 mg/kg per d for 7 d, per os) dose-dependently promoted NGF release in the mouse hippocampus, leading to phosphorylation of extracellular signal-regulated kinases and cyclic AMP response element-binding protein. ME significantly increased pre- and post-synapse formation, acetylcholine synthesisation, neuronal cell differentiation, neurite outgrowth and neuronal cell proliferation in the mouse hippocampus. Furthermore, ME significantly increased latency time in the passive avoidance task (P< 0·001) and recognition time of novel objects in the object recognition test (P< 0·05), indicating improvements in learning and memory. Taken together, these data suggest that ME exhibits a memory-enhancing effect via up-regulation of NGF.

  18. Solubilization of nerve growth factor receptors of rabbit superior cervical ganglia.

    PubMed

    Banerjee, S P; Cuatrecasas, P; Snyder, S H

    1976-09-25

    Nerve growth factor (NGF) receptors of rabbit superior cervical ganglia can be solubilized by treatment with detergents and readily assayed in the soluble state. Triton X-100 and deoxycholate reduce specific binding of NGF to ganglia membranes. In membranes treated with Triton X-100 (0.5 to 2.0%) the reduction in NGF binding by membranes is accompanied by a corresponding increase in binding in the supernatant fluid. NGF binding in soluble preparations can be rapidly assayed by precipitating NGF bound to receptors with polyethylene glycol under conditions in which unbound NGF is not precipitated. NGF binding to soluble preparations is saturable whether evaluated by the binding of 125I-NGF or by diluting 125I-NGF with native NGF. Using both techniques, the dissociation constant for NGF binding to soluble receptors is about 0.2 nM, the same as its dissociation constant from receptor sites in intact membranes. NGF binding to soluble receptors displays a high degree of peptide specificity, similar to receptor sites in intact membranes of superior cervical ganglia. A method of labeling NGF with 125I-3(4-hydroxyphenyl) propionic acid N-hydroxysuccinimide ester is described which leads to binding properties that are superior to those obtained with previously described 125I-NGF preparations.

  19. Locally applied nerve growth factor enhances bone consolidation in a rabbit model of mandibular distraction osteogenesis.

    PubMed

    Wang, Lei; Zhou, Shuxia; Liu, Baolin; Lei, Delin; Zhao, Yinghua; Lu, Chao; Tan, Aixing

    2006-12-01

    Distraction osteogenesis is widely used in treating deformities, defects, and fractures of both long bones and craniofacial bones. Demands for acceleration of bone consolidation are increased in distraction osteogenesis. Nerve growth factor (NGF) can enhance innervation and bone regeneration in a fracture model and stimulate differentiation of osteoblastic cells. In this study, we tested the ability of locally applied NGF to enhance bone regeneration in a rabbit model of mandibular distraction osteogenesis. Twenty rabbits underwent bilateral distraction osteogenesis with a rate of 0.5 mm per 12 h. Two times 0.04 mg human NGFbeta (hNGFbeta) in buffer was injected into the callus after distraction. The contralateral side received placebo injections. Rabbits were euthanized at consolidation times of 14 and 28 days. Specimens were subjected to radiography, callus dimensions measurement, mechanical testing, and bone histological and histomorphometric analysis. The maximum load, bone volume/total volume, mineral apposition rate of the 1st to 11th day, and mineralized bone percentage were significantly higher in the hNGFbeta side at 14 and 28 days (p<0.05). The data indicate that locally applied hNGFbeta can accelerate callus maturation and may be an option to shorten the consolidation period in distraction osteogenesis.

  20. Nerve growth factor receptor from rabbit sympathetic ganglia membranes. Relationship between subforms.

    PubMed

    Kouchalakos, R N; Bradshaw, R A

    1986-12-05

    The receptor for nerve growth factor (NGF) was purified from Triton X-100 extracts of sympathetic ganglia membranes by affinity chromatography on NGF-Sepharose. Elution of purified receptor was accomplished at pH 5 in the presence of 1 M NaCl. Sodium dodecyl sulfate gel electrophoresis of the purified iodinated receptor showed three major bands at Mr = 126,000, Mr = 105,000, and Mr = 81,000. Affinity labeling of the purified receptor using 125I-NGF and the photoreactive agent N-hydroxysuccinimidyl-p-azidobenzoate resulted in two major cross-linked complexes corresponding to Mr = 135,000 and Mr = 110,000. This labeling pattern is similar to that observed with sympathetic ganglia membranes (Massague, J., Guillette, B. J., Czech, M. P., Morgan, C. J., and Bradshaw, R. A. (1981) J. Biol. Chem. 256, 9419-9424) and indicates that these two forms do not arise from the cross-linking procedure. Reaction of the photoaffinity labeled NGF receptors with increasing amounts of trypsin resulted in a progressive decrease in the high molecular weight complex with a concomitant increase in the low molecular weight form. When the larger complex was isolated by electroelution from a sodium dodecyl sulfate gel and treated with trypsin, a species corresponding to Mr = 100,000 was generated. These observations are best explained by a precursor-product relationship for the two NGF receptor species of sympathetic neurons.

  1. Nerve growth factor and nociception: from experimental embryology to new analgesic therapy.

    PubMed

    Lewin, Gary R; Lechner, Stefan G; Smith, Ewan St John

    2014-01-01

    Nerve growth factor (NGF) is central to the development and functional regulation of sensory neurons that signal the first events that lead to pain. These sensory neurons, called nociceptors, require NGF in the early embryo to survive and also for their functional maturation. The long road from the discovery of NGF and its roles during development to the realization that NGF plays a major role in the pathophysiology of inflammatory pain will be reviewed. In particular, we will discuss the various signaling events initiated by NGF that lead to long-lasting thermal and mechanical hyperalgesia in animals and in man. It has been realized relatively recently that humanized function blocking antibodies directed against NGF show remarkably analgesic potency in human clinical trials for painful conditions as varied as osteoarthritis, lower back pain, and interstitial cystitis. Thus, anti-NGF medication has the potential to make a major impact on day-to-day chronic pain treatment in the near future. It is therefore all the more important to understand the precise pathways and mechanisms that are controlled by NGF to both initiate and sustain mechanical and thermal hyperalgesia. Recent work suggests that NGF-dependent regulation of the mechanosensory properties of sensory neurons that signal mechanical pain may open new mechanistic avenues to refine and exploit relevant molecular targets for novel analgesics.

  2. Nerve growth factor in the adult brain of a teleostean model for aging research: Nothobranchius furzeri.

    PubMed

    D'Angelo, L; Castaldo, L; Cellerino, A; de Girolamo, P; Lucini, C

    2014-07-01

    Nerve growth factor (NGF) acts on central nervous system neurons, regulating naturally occurring cell death, synaptic connectivity, fiber guidance and dendritic morphology. The dynamically regulated production of NGF beginning in development, extends throughout adult life and aging, exerting numerous roles through a surprising variety of neurons and glial cells. This study analyzes the localization of NGF in the brain of the teleost fish Nothobranchius furzeri, an emerging model for aging research due to its short lifespan. Immunochemical and immunohistochemical experiments were performed by employing an antibody mapping at the N-terminus of the mature chain human origin NGF. Western blot analysis revealed an intense and well defined band of 20 kDa, which corresponds to proNGF of N. furzeri. Immunohistochemistry revealed NGF immunoreactivity (IR) diffused throughout all regions of telencephalon, diencephalon, mesencephalon and rhomboencephalon. It was detected in neurons and in glial cells, the latter mostly lining the mesencephalic and rhomboencephalic ventricles. Particularly in neurons, NGF IR was localized in perikarya and, to a less extent, in fibers. The widespread distribution of proNGF suggests that it might modulate numerous physiological functions in the adult brain of N. furzeri. The present survey constitutes a baseline study to enhance the understanding of the mechanisms underlying the role of NGF during aging processes.

  3. Amplification and Temporal Filtering during Gradient Sensing by Nerve Growth Cones Probed with a Microfluidic Assay

    PubMed Central

    Morel, Mathieu; Shynkar, Vasyl; Galas, Jean-Christophe; Dupin, Isabelle; Bouzigues, Cedric; Studer, Vincent; Dahan, Maxime

    2012-01-01

    Nerve growth cones (GCs) are chemical sensors that convert graded extracellular cues into oriented axonal motion. To ensure a sensitive and robust response to directional signals in complex and dynamic chemical landscapes, GCs are presumably able to amplify and filter external information. How these processing tasks are performed remains however poorly known. Here, we probe the signal-processing capabilities of single GCs during γ-Aminobutyric acid (GABA) directional sensing with a shear-free microfluidic assay that enables systematic measurements of the GC output response to variable input gradients. By measuring at the single molecule level the polarization of GABAA chemoreceptors at the GC membrane, as a function of the external GABA gradient, we find that GCs act as i), signal amplifiers over a narrow range of concentrations, and ii), low-pass temporal filters with a cutoff frequency independent of stimuli conditions. With computational modeling, we determine that these systems-level properties arise at a molecular level from the saturable occupancy response and the lateral dynamics of GABAA receptors. PMID:23083707

  4. Reduced nerve growth factor levels in stress-related brain regions of folate-deficient mice.

    PubMed

    Eckart, S; Hörtnagl, H; Kronenberg, G; Gertz, K; Hörster, H; Endres, M; Hellweg, R

    2013-08-15

    Folate deficiency has been linked to neurodegenerative and stress-related diseases such as stroke, dementia and depression. The role of the neurotrophins nerve growth factor (NGF) and neurotrophin-3 (NT-3) in stress-related disorders and neurodegeneration has garnered increasing attention in recent years. Uracil misincorporation is involved in the neuropsychiatric dysfunction induced by experimental folate deprivation. However, the effects of folate deficiency on the expression of NGF and NT-3 in brain tissue have not yet been investigated. In a 2×2 design, aged mice lacking uracil-DNA N-glycosylase (Ung(-/-)) versus wild-type (Ung(+/+)) controls were subjected to a folate-deficient diet versus a regular diet for three months. Independent of genotype, folate deficiency led to decreased NGF protein levels in the frontal cortex and amygdala. In the hippocampus, NGF levels were increased in UNG(-/-) mice on the normal diet, but not under folate deficiency, while in UNG(+/+) mice, folate deprivation did not affect hippocampal NGF content. NT-3 protein concentrations were neither affected by genotype nor by folate deficiency. Altogether, the results of our study show that folate deficiency affects NGF levels in the frontal cortex, amygdala and hippocampus. The decrease in NGF content in the hippocampus in response to folate deficiency in Ung(-/-) mice may contribute to their phenotype of enhanced anxiety and despair-like behavior as well as to selective hippocampal neurodegeneration. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

  5. Amyloid beta-induced nerve growth factor dysmetabolism in Alzheimer disease.

    PubMed

    Bruno, Martin A; Leon, Wanda C; Fragoso, Gabriela; Mushynski, Walter E; Almazan, Guillermina; Cuello, A Claudio

    2009-08-01

    We previously reported that the precursor form of nerve growth factor (pro-NGF) and not mature NGF is liberated in the CNS in an activity-dependent manner, and that its maturation and degradation occur in the extracellular space by the coordinated action of proteases.Here, we present evidence of diminished conversion of pro-NGF to its mature form and of greater NGF degradation in Alzheimer disease (AD) brain samples compared with controls. These alterations of the NGF metabolic pathway likely resulted in the increased pro-NGF levels. The pro-NGF was largely in a peroxynitrited form in the AD samples. Intrahippocampal injection of amyloid-beta oligomers provoked similar upregulation of pro-NGF in naive rats that was accompanied by evidence of microglial activation (CD40), increased levels of inducible nitric oxide synthase, and increased activity of the NGF-degrading enzyme matrix metalloproteinase 9. The elevated inducible nitric oxide synthase provoked the generation of biologically inactive, peroxynitrite-modified pro-NGF in amyloid-beta oligomer-injected rats. These parameters were corrected by minocycline treatment. Minocycline also diminished altered matrix metalloproteinase 9, inducible nitric oxide synthase, and microglial activation (CD40); improved cognitive behavior; and normalized pro-NGF levels in a transgenic mouse AD model. The effects of amyloid-beta amyloid CNS burden on NGF metabolism may explain the paradoxical upregulation of pro-NGF in AD accompanied by atrophy of forebrain cholinergic neurons.

  6. Nerve growth factor plasma levels and ventricular repolarization in Rett syndrome.

    PubMed

    Guideri, F; Acampa, M; Calamandrei, G; Aloe, L; Zappella, M; Hayek, Y

    2004-01-01

    Rett syndrome is a severe neurological developmental disorder. In this syndrome, the high incidence of sudden death is correlated with an alteration of ventricular repolarization. The purpose of this study was to evaluate plasmatic levels of nerve growth factor (NGF) in Rett patients with prolonged corrected QT (QTc) interval in comparison with those of Rett patients with normal QTc. We observed 23 female Rett patients (9.9+/-4.7 years). NGF plasma levels and QTc interval were measured in all patients. Student t-test was performed for statistical analysis. NGF plasma levels were significantly lower in Rett patients with QTc interval prolongation (QTc > 0.44 sec) in comparison with Rett patients with a normal QTc interval (4.5+/-4.5 vs 11+/-8.3 pg/ml, p = 0.02). The alteration of NGF levels, observed in Rett patients with a long QTc interval, may explain the presence of an altered ventricular repolarization associated with a higher risk of cardiac arrhythmias.

  7. Enhanced bioavailability of nerve growth factor with phytantriol lipid-based crystalline nanoparticles in cochlea

    PubMed Central

    Bu, Meng; Tang, Jingling; Wei, Yinghui; Sun, Yanhui; Wang, Xinyu; Wu, Linhua; Liu, Hongzhuo

    2015-01-01

    Purpose Supplementation of exogenous nerve growth factor (NGF) into the cochlea of deafened animals rescues spiral ganglion cells from degeneration. However, a safe and potent delivery of therapeutic proteins, such as NGF, to spiral ganglion cells remains one of the greatest challenges. This study presents the development of self-assembled cubic lipid-based crystalline nanoparticles to enhance inner ear bioavailability of bioactive NGF via a round window membrane route. Methods A novel nanocarrier-entrapped NGF was developed based on phytantriol by a liquid precursor dilution, with Pluronic® F127 and propylene glycol as the surfactant and solubilizer, respectively. Upon dilution of the liquid lipid precursors, monodispersed submicron-sized particles with a slight negative charge formed spontaneously. Results Biological activity of entrapped NGF was assessed using pheochromocytoma cells with NGF-loaded reservoirs to induce significant neuronal outgrowth, similar to that seen in free NGF-treated controls. Finally, a 3.28-fold increase in inner ear bioavailability was observed after administration of phytantriol lipid-based crystalline nanoparticles as compared to free drug, contributing to an enhanced drug permeability of the round window membrane. Conclusion Data presented here demonstrate the potential of lipid-based crystalline nanoparticles to improve the outcomes of patients bearing cochlear implants. PMID:26604754

  8. Nerve growth factor amplifies cyclic AMP production in the HT4 neuronal cell line.

    PubMed

    Berg, K A; Maayani, S; McKay, R; Clarke, W P

    1995-01-01

    There has been considerable interest and controversy in the relationship between nerve growth factor (NGF) and the cyclic AMP (cAMP) second messenger system. We have used a novel, neuronal cell line (HT4) to investigate the effect of NGF on the adenylyl cyclase signaling system. Treatment of cells with NGF (100 ng/ml, 15 min) amplified cAMP accumulation (approximately 75%) in response to activation of adenosine A2 receptors (5 min) with 5'-N-ethylcarboxamidoadenosine or activation of adenylyl cyclase directly with forskolin. Basal cAMP accumulation was not altered by NGF. This amplification appears to be mediated by activation of protein kinase C (PKC) because (1) it was mimicked by activators (phorbol esters and a diacylglycerol analogue) of PKC, (2) the effects of NGF and phorbol ester on cAMP accumulation were not additive, (3) NGF amplification of cAMP accumulation was abolished by down-regulation of PKC, (4) NGF increased cytosolic PKC activity, and (5) inhibitors of PKC blocked the NGF-induced amplification of cAMP accumulation. Although NGF-induced amplification of cAMP accumulation was dependent upon PKC, mechanisms other than the classic activation pathway (i.e., hydrolysis of inositol phospholipids or the production of diacylglycerol) appeared to mediate PKC activation by NGF. The tyrosine kinase inhibitor, lavendustin A, blocked NGF-mediated amplification of cAMP accumulation, suggesting a novel interaction between a tyrosine kinase and protein kinase C.

  9. Nerve growth induces 5-HT sub 3 recognition sites in rat pheochromocytoma (PC12) cells

    SciTech Connect

    Gordon, J.C.; Rowland, H.C. )

    1990-01-01

    In rat pheochromocytoma (PC12) cells, nerve growth factor (7S NGF) induced the expression of recognition sites that bind the specific 5-HT{sub 3} antagonist (S-) ({sup 3}H) zacopride. Culturing PC12 cells for 8-12 days in the presence of 50 ng/ml NGF increased the density (B{sub max}) of (S-) ({sup 3}H) zacopride binding sites in cell membranes (0-100,000 x g fraction) from 0 to 105 fmoles/mg protein. This binding exhibited high affinity for (S-) ({sup 3}H) zacopride (K{sub d}=0.8 nM), was specific (>95%), and was inhibited by 5-HT{sub 3} compounds with a rank of potency (quipazine>ICS 205-930 > GR38032F > BRL 24924{approx}MDL 72222 > phenylbiguanide {le} seroton-in > 2-methyl-serotonin > metoclopramide) which was distinct from neuroblastoma cells. Thus, NGF-differentiated PC12 cells possess a 5-HT{sub 3} receptor and should be useful to investigate its regulation and biochemical mechanism of action.

  10. Structure-function studies of nerve growth factor: functional importance of highly conserved amino acid residues.

    PubMed Central

    Ibáñez, C F; Hallböök, F; Ebendal, T; Persson, H

    1990-01-01

    Selected amino acid residues in chicken nerve growth factor (NGF) were replaced by site-directed mutagenesis. Mutated NGF sequences were transiently expressed in COS cells and the yield of NGF protein in conditioned medium was quantified by Western blotting. Binding of each mutant to NGF receptors on PC12 cells was evaluated in a competition assay. The biological activity was determined by measuring stimulation of neurite outgrowth from chick sympathetic ganglia. The residues homologous to the proposed receptor binding site of insulin (Ser18, Met19, Val21, Asp23) were substituted by Ala. Replacement of Ser18, Met19 and Asp23 did not affect NGF activity. Modification of Val21 notably reduced both receptor binding and biological activity, suggesting that this residue is important to retain a fully active NGF. The highly conserved Tyr51 and Arg99 were converted into Phe and Lys respectively, without changing the biological properties of the molecule. However, binding and biological activity were greatly impaired after the simultaneous replacement of both Arg99 and Arg102 by Gly. The three conserved Trp residues at positions 20, 75 and 98 were substituted by Phe. The Trp mutated proteins retained 15-60% of receptor binding and 40-80% of biological activity, indicating that the Trp residues are not essential for NGF activity. However, replacement of Trp20 significantly reduced the amount of NGF in the medium, suggesting that this residue may be important for protein stability. Images Fig. 4. PMID:2328722

  11. Brain changes in Alzheimer's disease patients with implanted encapsulated cells releasing nerve growth factor.

    PubMed

    Ferreira, Daniel; Westman, Eric; Eyjolfsdottir, Helga; Almqvist, Per; Lind, Göran; Linderoth, Bengt; Seiger, Ake; Blennow, Kaj; Karami, Azadeh; Darreh-Shori, Taher; Wiberg, Maria; Simmons, Andrew; Wahlund, Lars-Olof; Wahlberg, Lars; Eriksdotter, Maria

    2015-01-01

    New therapies with disease-modifying effects are urgently needed for treating Alzheimer's disease (AD). Nerve growth factor (NGF) protein has demonstrated regenerative and neuroprotective effects on basal forebrain cholinergic neurons in animal studies. In addition, AD patients treated with NGF have previously shown improved cognition, EEG activity, nicotinic binding, and glucose metabolism. However, no study to date has analyzed brain atrophy in patients treated with NGF producing cells. In this study we present MRI results of the first clinical trial in patients with AD using encapsulated NGF biodelivery to the basal forebrain. Six AD patients received the treatment during twelve months. Patients were grouped as responders and non-responders according to their twelve-months change in MMSE. Normative values were created from 131 AD patients from ADNI, selecting 36 age- and MMSE-matched patients for interpreting the longitudinal changes in MMSE and brain atrophy. Results at baseline indicated that responders showed better clinical status and less pathological levels of cerebrospinal fluid (CSF) Aβ1-42. However, they showed more brain atrophy, and neuronal degeneration as evidenced by higher CSF levels of T-tau and neurofilaments. At follow-up, responders showed less brain shrinkage and better progression in the clinical variables and CSF biomarkers. Noteworthy, two responders showed less brain shrinkage than the normative ADNI group. These results together with previous evidence supports the idea that encapsulated biodelivery of NGF might have the potential to become a new treatment strategy for AD with both symptomatic and disease-modifying effects.

  12. Nerve growth factor for Bell’s palsy: A meta-analysis

    PubMed Central

    SU, YIPENG; DONG, XIAOMENG; LIU, JUAN; HU, YAOZHI; CHEN, JINBO

    2015-01-01

    A meta-analysis was performed to evaluate the efficacy and safety of nerve growth factor (NGF) in the treatment of Bell’s palsy. PubMed, the Cochrane Central Register of Controlled Trials, Embase and a number of Chinese databases, including the China National Knowledge Infrastructure, China Biology Medicine disc, VIP Database for Chinese Technical Periodicals and Wan Fang Data, were used to collect randomised controlled trials (RCTs) of NGF for Bell’s palsy. The span of the search covered data from the date of database establishment until December 2013. The included trials were screened comprehensively and rigorously. The efficacies of NGF were pooled via meta-analysis performed using Review Manager 5.2 software. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated using the fixed-effects model. The meta-analysis of eight RCTs showed favorable effects of NGF on the disease response rate (n=642; OR, 3.87; 95% CI, 2.13–7.03; P<0.01; I2=0%). However, evidence supporting the effectiveness of NGF for the treatment of Bell’s palsy is limited. The number and quality of trials are too low to form solid conclusions. Further meticulous RCTs are required to overcome the limitations identified in the present study. PMID:25574223

  13. Nerve growth factor protects against palmitic acid-induced injury in retinal ganglion cells

    PubMed Central

    Yan, Pan-shi; Tang, Shu; Zhang, Hai-feng; Guo, Yuan-yuan; Zeng, Zhi-wen; Wen, Qiang

    2016-01-01

    Accumulating evidence supports an important role for nerve growth factor (NGF) in diabetic retinopathy. We hypothesized that NGF has a protective effect on rat retinal ganglion RGC-5 cells injured by palmitic acid (PA), a metabolic factor implicated in the development of diabetes and its complications. Our results show that PA exposure caused apoptosis of RGC-5 cells, while NGF protected against PA insult in a concentration-dependent manner. Additionally, NGF significantly attenuated the levels of reactive oxygen species (ROS) and malondialdehyde (MDA) in RGC-5 cells. Pathway inhibitor tests showed that the protective effect of NGF was completely reversed by LY294002 (PI3K inhibitor), Akt VIII inhibitor, and PD98059 (ERK1/2 inhibitor). Western blot analysis revealed that NGF induced the phosphorylation of Akt/FoxO1 and ERK1/2 and reversed the PA-evoked reduction in the levels of these proteins. These results indicate that NGF protects RGC-5 cells against PA-induced injury through anti-oxidation and inhibition of apoptosis by modulation of the PI3K/Akt and ERK1/2 signaling pathways. PMID:28123432

  14. The interactions between nerve growth factor and gonadotrophins in bovine oviduct.

    PubMed

    Li, Chunjin; Ma, Yonghe; Yi, Kangle; Wang, Chunqiang; Li, Wanhong; Liu, Zhuo; Sun, Lina; Chen, Shuxiong; Yu, Jiaxin; Li, Hongjiao; Chen, Lu; Zhou, Xu

    2014-10-01

    Nerve growth factor promotes the survival and differentiation of nervous cells and is thought to play an important role in the development of reproductive tissues. The aims of this work were to detect the presence of NGF and its receptor NTRK1 in bovine oviduct samples, and to investigate the regulatory interactions between NGF/NTRK1 and gonadotrophins in bovine oviduct epithelial cells. Both transcripts and proteins of NGF and NTRK1 were detected by RT-PCR and Western blotting, and the corresponding proteins were specifically immunolocalized in oviduct epithelial cells. In addition, real-time PCR experiments revealed that the levels of NGF and NTRK1 mRNA in oviduct epithelial cells treated with exogenous FSH or LH were greater than those in negative control cells (P<0.05). Similarly, treatment with NGF significantly increased the expression of FSHR and LHR in oviduct epithelial cells via its effects on NTRK1 (P<0.05). This process was suppressed by treatment with the NTRK1 inhibitor K252α. We conclude that NGF/NTRK1 may have a role in regulating the function of bovine oviducts via its interactions with gonadotrophins.

  15. Methamphetamine reversed maternal separation-induced decrease in nerve growth factor in the ventral hippocampus.

    PubMed

    Dimatelis, J J; Russell, V A; Stein, D J; Daniels, W M

    2014-06-01

    Stress has been suggested to predispose individuals to drug abuse. The early life stress of maternal separation (MS) is known to alter the response to drugs of abuse later in life. Exposure to either stress or methamphetamine has been shown to alter neurotrophic factors in the brain. Changes in neurotrophin levels may contribute to the underlying molecular mechanisms responsible for drug use- and stress-induced behaviours. The purpose of the present study was to investigate the individual effects of MS and methamphetamine administration during adolescence and the combined effects of both stressors on brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) levels in the dorsal and ventral hippocampus (HC) in adulthood. Methamphetamine administration (1 mg/kg, daily from postnatal day (PND) 33 to 36 and from PND 39 to 42), MS and the combination of the two stressors resulted in decreased BDNF levels in both the dorsal and ventral HC. MS decreased NGF levels in the ventral HC which was restored by methamphetamine administration in adolescence. In the dorsal HC, NGF remained unaltered by either stressor alone or in combination. We propose that the restoration of NGF levels in the ventral HC may reflect a possible compensatory mechanism in response to methamphetamine exposure in adolescence following the early life stress of MS.

  16. Circulating nerve growth factor in primary and secondary Raynaud's syndrome - results of a pilot study.

    PubMed

    Klein-Weigel, P; Gutsche-Petrak, B; Humpel, C; Riemekasten, G; Ivanov, S; Heidrich, H

    2009-02-01

    In this pilot study we examined circulating concentrations of nerve growth factor (NGF) in controls and patients suffering from primary or secondary Raynaud's syndrome and determined their relation to digital vasospasm. Eighteen controls, 16 patients with primary RP and 19 patients with systemic sclerosis (SScl) were included. Degree of vasospasm was graduated according to the degree of plethysmographically measured vasospastic reaction after cold test. A diary was handed out for documentation of the daily number and duration of RP during a period of 16 days. Circulating NGF levels were analysed by a commercial ELISA (Promega Inc., USA). SScl-patients were significantly older (p < 0.0001) and more severely affected by spontaneously occurring RP (p = 0.03), whereas the severity of the vasospastic reactions after a standard cold test were not significantly different between the groups (p = 0.09). Within each study group and between the study groups elevated NGF levels were observed only in SScl-patients after thermal provocations (p = < 0.05). In a correlation analysis restricted to patients with PRP or SRP, the degree of vasospasm after cold testing as well as the intensity of Raynaud's symptoms were not correlated with NGF-levels (p = n.s.). Our results do not support the hypothesis that NGF plays a major role in the generation of vasospasm in Raynaud's phenomenon.

  17. Response Growth With Sound Level in Auditory-Nerve Fibers After Noise-Induced Hearing Loss

    PubMed Central

    Heinz, Michael G.; Young, Eric D.

    2010-01-01

    People with sensorineural hearing loss are often constrained by a reduced acoustic dynamic range associated with loudness recruitment; however, the neural correlates of loudness and recruitment are still not well understood. The growth of auditory-nerve (AN) activity with sound level was compared in normal-hearing cats and in cats with a noise-induced hearing loss to test the hypothesis that AN-fiber rate-level functions are steeper in impaired ears. Stimuli included best-frequency and fixed-frequency tones, broadband noise, and a brief speech token. Three types of impaired responses were observed. 1) Fibers with rate-level functions that were similar across all stimuli typically had broad tuning, consistent with outer-hair-cell (OHC) damage. 2) Fibers with a wide dynamic range and shallow slope above threshold often retained sharp tuning, consistent with primarily inner-hair-cell (IHC) damage. 3) Fibers with very steep rate-level functions for all stimuli had thresholds above approximately 80 dB SPL and very broad tuning, consistent with severe IHC and OHC damage. Impaired rate-level slopes were on average shallower than normal for tones, and were steeper in only limited conditions. There was less variation in rate-level slopes across stimuli in impaired fibers, presumably attributable to the lack of suppression-induced reductions in slopes for complex stimuli relative to BF-tone slopes. Sloping saturation was observed less often in impaired fibers. These results illustrate that AN fibers do not provide a simple representation of the basilar-membrane I/O function and suggest that both OHC and IHC damage can affect AN response growth. PMID:14534289

  18. Pro-nerve growth factor in the ovary and human granulosa cells

    PubMed Central

    Meinel, Sabine; Blohberger, Jan; Berg, Dieter; Berg, Ulrike; Dissen, Gregory A.; Ojeda, Sergio R.; Mayerhofer, Artur

    2016-01-01

    Background Pro-nerve growth factor must be cleaved to generate mature NGF, which was suggested to be a factor involved in ovarian physiology and pathology. Extracellular proNGF can induce cell death in many tissues. Whether extracellular proNGF exists in the ovary and may play a role in the death of follicular cells or atresia was unknown. Material and Methods Immunohistochemistry of human and Rhesus monkey ovarian sections was performed. IVF-derived follicular fluid and human granulosa cells were studied by RT-PCR, qPCR, Western blotting, ATP- and caspase-assays. Results and Conclusions Immunohistochemistry of ovarian sections identified proNGF in granulosa cells and Western blotting of human isolated granulosa cells confirmed the presence of proNGF. Ovarian granulosa cells thus produce proNGF. Recombinant human proNGF even at high concentrations did not affect the levels of ATP or the activity of caspase 3/7, indicating that in granulosa cells proNGF does not induce death. In contrast, mature NGF, which was detected previously in follicular fluid, may be a trophic molecule for granulosa cells with unexpected functions. We found that in contrast to proNGF, NGF increased the levels of the transcription factor early growth response 1 and of the enzyme choline acetyl-transferase. A mechanism for the generation of mature NGF from proNGF in the follicular fluid may be extracellular enzymatic cleavage. The enzyme MMP7 is known to cleave proNGF and was identified in follicular fluid and as a product of granulosa cells. Thus the generation of NGF in the ovarian follicle may depend on MMP7. PMID:26457789

  19. Germ cell mitogenic activity is associated with nerve growth factor-like protein(s).

    PubMed

    Onoda, M; Pflug, B; Djakiew, D

    1991-12-01

    The mitogenicity of germ cell proteins released from round spermatids (RS) and pachytene spermatocytes (PS) was investigated. Germ cells were isolated by centrifugal elutriation from 90-day-old rat testes and incubated in a supplement enriched culture media that lacked exogenous proteins. The conditioned culture media of RS and PS were dialysed/concentrated and lyophilized to prepare RS protein (RSP) and PS protein (PSP). Mitogenic activity of RSP and PSP was determined by 3H-thymidine incorporation into Swiss 3T3 fibroblasts. RSP and PSP stimulated 3H-thymidine incorporation by fibroblasts in a dose-dependent manner. At a higher concentration of RSP (300 micrograms/ml), fibroblast proliferation was stimulated from 6- to 20-fold of control cultures, whereas PSP (300 micrograms/ml) stimulated fibroblast proliferation 2.5-fold of control cultures. Since RSP exhibited substantially greater mitogenic activity than PSP we further investigated the RSP mitogenic substance(s) by immunoneutralization with antibodies against several growth factors. The mitogenic activity of RSP was significantly reduced by treatment with nerve growth factor (NGF) antibody, while neither the treatment of RSP with acidic fibroblast growth factor (aFGF) antibody, nor basic fibroblast growth factor (bFGF) antibody significantly modified the mitogenic activity of RSP. Interestingly, murine NGF-beta, recombinant human NGF-beta, and bovine serum albumin (BSA) did not exhibit mitogenic activity on 3T3 fibroblasts. Nevertheless, the presence of a NGF-like protein in RS and PS was confirmed by indirect immunofluorescence staining with a murine NGF antibody. Subsequently, a Western blot analysis with the NGF antibody identified two immunoreactive bands of 41 +/- 2 kDa and 51 +/- 1 kDa in both RSP and PSP under reduced conditions. These germ cell NGF-like proteins were apparently different from similarly prepared murine and human NGFs (13 kDa) in their molecular weight. Furthermore, neurite outgrowth

  20. Nerve growth factor (NGF) promotes angiogenesis in the quail chorioallantoic membrane.

    PubMed

    Lazarovici, Philip; Gazit, Aviv; Staniszewska, Izabela; Marcinkiewicz, Cezary; Lelkes, Peter I

    2006-01-01

    Angiogenesis, the formation of new blood vessels, is tightly regulated by growth factors, such as vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF). The authors hypothesize that nerve growth factor (NGF), a well known neurotrophin, may play a direct angiogenic role. To test this hypothesis, the authors measured the effects of NGF on the natural vascularization of the quail chorioallantoic membrane (CAM). The angiogenic effect of NGF was compared to that of human recombinant VEGF165 (rhVEGF) and basic FGF (rhbFGF). In comparison to phosphate-buffered saline-treated controls, NGFs from different biological sources (mouse, viper, and cobra) increased the rate of angiogenesis in a dose-dependent fashion from 0.5 to 5 microg. For quantitative morphometry, grayscale images of the blood vessels end points of the CAM arteries were binarized for visualization and skeletonized for quantization by fractal analysis. In mouse NGF-treated embryos the fractal dimension (Df), indicative of arterial vessel length and density, increased to 1.266 +/- 0.021 compared to 1.131 +/- 0.018 (p < .001) for control embryos. This effect was similar to that of 0.5 microg rhVEGF (1.290 +/- 0.021, p < .001) and 1.5 microg rhbFGF (1.264 +/- 0.028, p < .001). The mouse NGF-induced angiogenic effect was blocked by 1 microM K252a (1.149 +/- 0.018, p < .001), an antagonist of the NGF/trkA receptor, but not by 1 microM SU-5416 (1.263 +/- 0.029, p < .001), the VEGF/Flk1 receptor antagonist, indicating a direct, selective angiogenic effect of NGF via quail embryo trkA receptor activation. These results confirm previous observations that NGF has angiogenic activity and suggest that this neurotrophin may also play an important role in the cardiovascular system, besides its well-known effects in the nervous system. The angiogenic properties of NGF may be beneficial in engineering new blood vessels and for developing novel antiangiogenesis therapies for cancer.

  1. Hepatocyte growth factor, vascular endothelial growth factor, glial cell-derived neurotrophic factor and nerve growth factor are differentially affected by early chronic ethanol or red wine intake.

    PubMed

    Fiore, Marco; Mancinelli, Rosanna; Aloe, Luigi; Laviola, Giovanni; Sornelli, Federica; Vitali, Mario; Ceccanti, Mauro

    2009-08-10

    Ethanol intake during pregnancy and lactation induces severe changes in brain and liver throughout mechanisms involving growth factors. These are signaling molecules regulating survival, differentiation, maintenance and connectivity of brain and liver cells. Ethanol is an element of red wine which contains also compounds with antioxidant properties. Aim of the study was to investigate differences in hepatocyte growth factor (HGF), vascular endothelial growth factor (VEGF), glial cell-derived neurotrophic factor (GDNF) and nerve growth factor (NGF) in brain areas and liver by ELISA of 1-month-old male mice exposed perinatally to ethanol at 11 vol.% or to red wine at same ethanol concentration. Ethanol was administered before and during pregnancy up to pups' weaning. Ethanol per se elevated HGF in liver and cortex, potentiated liver VEGF, reduced GDNF in the liver and decreased NGF content in hippocampus and cortex in the offspring. We did not find changes in HGF or NGF due to red wine exposure. However, we revealed elevation in VEGF levels in liver and reduced GDNF in the cortex of animals exposed to red wine but the VEGF liver increase was more marked in animals exposed to ethanol only compared to the red wine group. In conclusion the present findings in the mouse show differences in ethanol-induced toxicity when ethanol is administered alone or in red wine that may be related to compounds with antioxidant properties present in the red wine.

  2. Effect of administration of antibodies against nerve growth factor in a rat model of muscle injury.

    PubMed

    Suzuki, Masahiro; Inage, Kazuhide; Sakuma, Yoshihiro; Orita, Sumihisa; Yamauchi, Kazuyo; Suzuki, Takane; Suzuki, Miyako; Kubota, Go; Oikawa, Yasuhiro; Sainoh, Takeshi; Sato, Jun; Fujimoto, Kazuki; Shiga, Yasuhiro; Abe, Koki; Kanamoto, Hirohito; Takahashi, Kazuhisa; Ohtori, Seiji

    2016-03-01

    Although muscle injury is a common source of pain, the mechanism causing such pain is not completely known. We have previously reported nerve growth factor (NGF) as a proinflammatory mediator involved in acute pain, and clinical trials have shown the effectiveness of anti-NGF antibodies for management of low back pain. Here, we aim to examine the effects of anti-NGF antibodies on muscle-derived pain by studying their effects on sensory innervation in a rat muscle injury model. A nervous system tracer, Fluoro-Gold, was applied to both gastrocnemius muscles of 24 male Sprague Dawley rats to stain the sensory nerves. Then, the drop-mass method was used to damage the right gastrocnemius muscle of the posterior limb. Anti-NGF antibodies (50μL) were injected into the injured muscles in 12 rats. Tissues were evaluated 1, 3, and 7 days post-injury by performing haematoxylin-and-eosin (HE) staining. The percentage of the total number of FG-positive cells that were also positive for a pain-related neuropeptide, calcitonin gene-related peptide (CGRP), was determined for the bilateral dorsal root ganglia from L1 to L6 7 days post-injury. HE staining showed active inflammation, indicated by increased basophil and eosinophil accumulation, at the injury site 1 and 3 days post-injury, as well as scar tissue formation 7 days post-injury. Injection of anti-NGF reduced muscle necrosis 1 and 3 days post-injury, and resulted in replacement of granulation tissue and muscle fibre regeneration 7 days post-injury. Anti-NGF also significantly inhibited CGRP among FG-positive cells (treatment group 38.2%, control group 49.6%; P<0.05). This study found active inflammation induced by NGF, which may contribute to pain after muscle injury. Anti-NGF antibodies successfully suppressed the pain mediator NGF and inhibited inflammation, suggesting NGF as a target for control in pain management. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Nerve growth factor regulates the expression of bradykinin binding sites on adult sensory neurons via the neurotrophin receptor p75.

    PubMed

    Petersen, M; Segond von Banchet, G; Heppelmann, B; Koltzenburg, M

    1998-03-01

    Neurotrophins mediate specific effects on sensory neurons through tyrosine kinase receptors. Most of these neurons also co-express the neurotrophin receptor p75 (p75NTR), but its function has remained obscure. We now show that nerve growth factor but not brain-derived neurotrophic factor or neurotrophin-3 selectively increases the expression of bradykinin binding sites on cultured dorsal root ganglion neurons from adult mouse via p75NTR. This up-regulation of bradykinin binding sites did not occur in neurons from mice lacking p75NTR or in neurons from wild-type mice treated with p75NTR-blocking antibody, indicating that tyrosine kinase receptors alone are not sufficient to trigger this physiological neuronal response. Thus, the interaction of nerve growth factor with p75NTR is an important factor contributing to chronic pain conditions.

  4. Clues from Crouzon: Insights into the potential role of growth factors in the pathogenesis of myelinated retinal nerve fibers.

    PubMed

    Garcia, Giancarlo A; Tian, Jack J; Apinyawasisuk, Supanut; Kim, Sarah; Akil, Handan; Sadun, Alfredo A

    2016-12-01

    We present a case of bilateral extensive peripapillary myelinated retinal nerve fibers (MRNF) in an individual with Crouzon syndrome, an inherited form of craniosynostosis caused by overactivation of fibroblast growth factor receptor 2. As a secondary aim, we examine the utility of optical coherence tomography (OCT) angiography for visualization of peripapillary vasculature obscured by myelination on other imaging modalities. A 24-year-old woman with Crouzon syndrome was evaluated for suspected optic neuritis in the right eye. Funduscopic examination and photography revealed the incidental finding of bilateral extensive peripapillary MRNF. OCT angiography provided excellent visualization of peripapillary retinal vessels, which were partially obscured by myelination on other imaging modalities. This association of Crouzon syndrome with bilateral peripapillary MRNF may lend insight into the developmental control of optic nerve myelination, the pathogenesis of MRNF, and the potential role of growth factors in these processes. Further, OCT angiography allowed for excellent blood vessel visualization in this case of MRNF.

  5. The topical application of nerve growth factor as a pharmacological tool for human corneal and skin ulcers.

    PubMed

    Aloe, Luigi; Tirassa, Paola; Lambiase, Alessandro

    2008-04-01

    The nerve growth factor is a soluble protein produced by and acting upon a number of different cells located in the nervous, endocrine and immune systems. Recent studies have shown that nerve growth factor (NGF) exerts a critical role on epithelial cells and fibroblasts under normal and pathological conditions. In this review, we present data prospecting the clinical potentiality of NGF in cutaneous and ocular "non-healing" chronic ulcers. A consistent number of in vitro and in vivo studies carried out on animal models and in humans indicated that fibroblasts and epithelial cells are receptive to the action of NGF and that NGF promotes skin and cornea ulcer healing. These observations lead to the hypothesis that NGF can be a potential useful pharmacological agent for clinical investigations. The available clinical evidences suggest that the topical application of NGF promotes healing action without side effects on corneal and cutaneous tissues damaged by chemical, physical and surgical insults and autoimmune disorders.

  6. Nerve growth factor injected into the gastric ulcer base incorporates into endothelial, neuronal, glial and epithelial cells: implications for angiogenesis, mucosal regeneration and ulcer healing.

    PubMed

    Tanigawa, T; Ahluwalia, A; Watanabe, T; Arakawa, T; Tarnawski, A S

    2015-08-01

    A previous study has demonstrated that locally administered growth factors such as epidermal growth factor, basic fibroblast growth factor and hepatocyte growth factor can accelerate healing of experimental gastric ulcers in rats. That study indicates that locally administered growth factors can exert potent biological effects resulting in enhanced gastric ulcers healing. However, the fate of injected growth factors, their retention and localization to specific cellular compartments have not been examined. In our preliminary study, we demonstrated that local injection of nerve growth factor to the base of experimental gastric ulcers dramatically accelerates ulcer healing, increases angiogenesis - new blood vessel formation, and improves the quality of vascular and epithelial regeneration. Before embarking on larger, definitive and time sequence studies, we wished to determine whether locally injected nerve growth factor is retained in gastric ulcer's tissues and taken up by specific cells during gastric ulcer healing. Gastric ulcers were induced in anesthetized rats by local application of acetic acid using standard methods; and, 60 min later fluorescein isothiocyanate-labeled nerve growth factor was injected locally to the ulcer base. Rats were euthanized 2, 5 and 10 days later. Gastric specimens were obtained and processed for histology. Unstained paraffin sections were examined under a fluorescence microscope, and the incorporation of fluorescein isothiocyanate-labeled nerve growth factor into various gastric tissue cells was determined and quantified. In addition, we performed immunostaining for S100β protein that is expressed in neural components. Five and ten days after ulcer induction labeled nerve growth factor (injected to the gastric ulcer base) was incorporated into endothelial cells of blood vessels, neuronal, glial and epithelial cells, myofibroblasts and muscle cells. This study demonstrates for the first time that during gastric ulcer healing

  7. Nerve growth factor and cytokines mediate lymphoid tissue-induced neurite outgrowth from mouse superior cervical ganglia in vitro.

    PubMed

    Kannan, Y; Bienenstock, J; Ohta, M; Stanisz, A M; Stead, R H

    1996-07-01

    Superior cervical ganglia (SCG) from neonatal mice were cultured with adult murine lymphoid tissue explants in Matrigel (Collaborative Biomedical, Bedford, MA). After 1 and 2 days in culture, many neurites grew toward thymus and spleen. Normal mesenteric lymph node (MLN) induced a smaller effect; however, activated MLN (isolated from mice 10 days after infection with Nippostrongylus brasiliensis; Nb-MLN-10d) caused significantly increased neurite outgrowth. To determine the roles of nerve growth factor (NGF) and cytokines in the promotion of neuritogenesis by lymphoid tissues, anti-NGF and various anti-cytokines were added to cocultures. Anti-NGF inhibited most of the neurite outgrowth toward thymus and spleen but only partially that toward Nb-MLN-10d. Anti-mouse IL-1 beta also significantly reduced the number of neurites growing toward thymus, spleen, and normal MLN. The number of neurites growing toward Nb-MLN-10d was significantly reduced by anti-IL-1 beta, anti-IL-3, anti-IL-6, or anti-GM-CSF. Exogenous IL-1 beta and IL-3 caused neurite outgrowth in single SCG cultures; and the IL-1 beta-, but not the IL-3-, mediated effect was completely blocked by anti-NGF. In one-day thymus/SCG cocultures, endogenous IL-1 was not detectable at concentrations sufficient to cause nerve growth; however, ample NGF was present in the thymic tissues and culture supernatants, but not in SCG. These data suggest that IL-1 mediates NGF production in lymphoid tissues, which in turn induces the growth of sympathetic nerves. Moreover, IL-3, IL-6, or GM-CSF produced during inflammation might also play important roles in the stimulation of nerve growth in vivo.

  8. [Construction of recombinant human nerve growth factor (rh-β-NGF) eukaryotic vector and its expression in HEK293 cells].

    PubMed

    Li, Jingchuan; Xue, Bofu; Yuan, Yuan; Ma, Mo; Zhu, Lin; Milburn, Rebecca; Le, Li; Hu, Peizhen; Ye, Jing

    2015-03-01

    Human nerve growth factor (NGF) is a nerve cell growth regulation factor, which can provide nutrition for the neurons and promote the neurites outgrowth. In order to produce large-scale recombinant human nerve growth factor (rh-beta-NGF), we constructed a plasmid vector, which can stably express the rh-beta-NGF in the HEK293 cell lines. First, the plasmid of pCMV-beta-NGF-IRES-dhfr was constructed and transformed into HEK293 cells. Then MTX pressurized filter and limiting dilution methods were used to obtain monoclonal HEK293 cell lines. After stepwise reducing serum in culture media, the cells eventually adapted to serum-free medium and secreted rh-beta-NGF. SDS-PAGE analysis revealed that the expression product owned a molecular weight of about 13 kDa and a purity of more than 50%. The peptide mapping sequencing analysis demonstrated the sequences of rh-beta-NGF matched with the theoretical ones. Later we purified this protein by ion exchange and molecular sieve chromatograph. Finally, our experimental results exhibited that the recombinant cell lines can stably express rh-beta-NGF with a high efficiency of more than 20 pg/cell x day. In addition, this protein could successfully induce differentiation of PC12 cells. In summary, our recombinant HEK293 cells can express bio-active rh-beta-NGF with great efficiency and stability, which supply a valid basis to large-scale production of rh-beta-NGF.

  9. Axonal Growth Arrests After an Increased Accumulation of Schwann Cells Expressing Senescence Markers and Stromal Cells in Acellular Nerve Allografts.

    PubMed

    Poppler, Louis H; Ee, Xueping; Schellhardt, Lauren; Hoben, Gwendolyn M; Pan, Deng; Hunter, Daniel A; Yan, Ying; Moore, Amy M; Snyder-Warwick, Alison K; Stewart, Sheila A; Mackinnon, Susan E; Wood, Matthew D

    2016-07-01

    Acellular nerve allografts (ANAs) and other nerve constructs do not reliably facilitate axonal regeneration across long defects (>3 cm). Causes for this deficiency are poorly understood. In this study, we determined what cells are present within ANAs before axonal growth arrest in nerve constructs and if these cells express markers of cellular stress and senescence. Using the Thy1-GFP rat and serial imaging, we identified the time and location of axonal growth arrest in long (6 cm) ANAs. Axonal growth halted within long ANAs by 4 weeks, while axons successfully regenerated across short (3 cm) ANAs. Cellular populations and markers of senescence were determined using immunohistochemistry, histology, and senescence-associated β-galactosidase staining. Both short and long ANAs were robustly repopulated with Schwann cells (SCs) and stromal cells by 2 weeks. Schwann cells (S100β(+)) represented the majority of cells repopulating both ANAs. Overall, both ANAs demonstrated similar cellular populations with the exception of increased stromal cells (fibronectin(+)/S100β(-)/CD68(-) cells) in long ANAs. Characterization of ANAs for markers of cellular senescence revealed that long ANAs accumulated much greater levels of senescence markers and a greater percentage of Schwann cells expressing the senescence marker p16 compared to short ANAs. To establish the impact of the long ANA environment on axonal regeneration, short ANAs (2 cm) that would normally support axonal regeneration were generated from long ANAs near the time of axonal growth arrest ("stressed" ANAs). These stressed ANAs contained mainly S100β(+)/p16(+) cells and markedly reduced axonal regeneration. In additional experiments, removal of the distal portion (4 cm) of long ANAs near the time of axonal growth arrest and replacement with long isografts (4 cm) rescued axonal regeneration across the defect. Neuronal culture derived from nerve following axonal growth arrest in long ANAs revealed no

  10. Axonal Growth Arrests After an Increased Accumulation of Schwann Cells Expressing Senescence Markers and Stromal Cells in Acellular Nerve Allografts

    PubMed Central

    Poppler, Louis H.; Ee, Xueping; Schellhardt, Lauren; Hoben, Gwendolyn M.; Pan, Deng; Hunter, Daniel A.; Yan, Ying; Moore, Amy M.; Snyder-Warwick, Alison K.; Stewart, Sheila A.; Mackinnon, Susan E.

    2016-01-01

    Acellular nerve allografts (ANAs) and other nerve constructs do not reliably facilitate axonal regeneration across long defects (>3 cm). Causes for this deficiency are poorly understood. In this study, we determined what cells are present within ANAs before axonal growth arrest in nerve constructs and if these cells express markers of cellular stress and senescence. Using the Thy1-GFP rat and serial imaging, we identified the time and location of axonal growth arrest in long (6 cm) ANAs. Axonal growth halted within long ANAs by 4 weeks, while axons successfully regenerated across short (3 cm) ANAs. Cellular populations and markers of senescence were determined using immunohistochemistry, histology, and senescence-associated β-galactosidase staining. Both short and long ANAs were robustly repopulated with Schwann cells (SCs) and stromal cells by 2 weeks. Schwann cells (S100β+) represented the majority of cells repopulating both ANAs. Overall, both ANAs demonstrated similar cellular populations with the exception of increased stromal cells (fibronectin+/S100β−/CD68− cells) in long ANAs. Characterization of ANAs for markers of cellular senescence revealed that long ANAs accumulated much greater levels of senescence markers and a greater percentage of Schwann cells expressing the senescence marker p16 compared to short ANAs. To establish the impact of the long ANA environment on axonal regeneration, short ANAs (2 cm) that would normally support axonal regeneration were generated from long ANAs near the time of axonal growth arrest (“stressed” ANAs). These stressed ANAs contained mainly S100β+/p16+ cells and markedly reduced axonal regeneration. In additional experiments, removal of the distal portion (4 cm) of long ANAs near the time of axonal growth arrest and replacement with long isografts (4 cm) rescued axonal regeneration across the defect. Neuronal culture derived from nerve following axonal growth arrest in long ANAs revealed no deficits

  11. Nerve growth cone lamellipodia contain two populations of actin filaments that differ in organization and polarity

    PubMed Central

    1992-01-01

    The organization and polarity of actin filaments in neuronal growth cones was studied with negative stain and freeze-etch EM using a permeabilization protocol that caused little detectable change in morphology when cultured nerve growth cones were observed by video- enhanced differential interference contrast microscopy. The lamellipodial actin cytoskeleton was composed of two distinct subpopulations: a population of 40-100-nm-wide filament bundles radiated from the leading edge, and a second population of branching short filaments filled the volume between the dorsal and ventral membrane surfaces. Together, the two populations formed the three- dimensional structural network seen within expanding lamellipodia. Interaction of the actin filaments with the ventral membrane surface occurred along the length of the filaments via membrane associated proteins. The long bundled filament population was primarily involved in these interactions. The filament tips of either population appeared to interact with the membrane only at the leading edge; this interaction was mediated by a globular Triton-insoluble material. Actin filament polarity was determined by decoration with myosin S1 or heavy meromyosin. Previous reports have suggested that the polarity of the actin filaments in motile cells is uniform, with the barbed ends toward the leading edge. We observed that the actin filament polarity within growth cone lamellipodia is not uniform; although the predominant orientation was with the barbed end toward the leading edge (47-56%), 22-25% of the filaments had the opposite orientation with their pointed ends toward the leading edge, and 19-31% ran parallel to the leading edge. The two actin filament populations display distinct polarity profiles: the longer filaments appear to be oriented predominantly with their barbed ends toward the leading edge, whereas the short filaments appear to be randomly oriented. The different length, organization and polarity of the two filament

  12. Atropine-enhanced, antigen challenge-induced airway hyperreactivity in guinea pigs is mediated by eosinophils and nerve growth factor.

    PubMed

    Verbout, Norah G; Jacoby, David B; Gleich, Gerald J; Fryer, Allison D

    2009-08-01

    Although anticholinergic therapy inhibits bronchoconstriction in asthmatic patients and antigen-challenged animals, administration of atropine 1 h before antigen challenge significantly potentiates airway hyperreactivity and eosinophil activation measured 24 h later. This potentiation in airway hyperreactivity is related to increased eosinophil activation and is mediated at the level of the airway nerves. Since eosinophils produce nerve growth factor (NGF), which is known to play a role in antigen-induced airway hyperreactivity, we tested whether NGF mediates atropine-enhanced, antigen challenge-induced hyperreactivity. Antibody to NGF (Ab NGF) was administered to sensitized guinea pigs with and without atropine pretreatment (1 mg/kg iv) 1 h before challenge. At 24 h after challenge, animals were anesthetized, vagotomized, paralyzed, and ventilated. Electrical stimulation of both vagus nerves caused bronchoconstriction that was increased in challenged animals. Atropine pretreatment potentiated antigen challenge-induced hyperreactivity. Ab NGF did not affect eosinophils or inflammatory cells in any group, nor did it prevent hyperreactivity in challenged animals that were not pretreated with atropine. However, Ab NGF did prevent atropine-enhanced, antigen challenge-induced hyperreactivity and eosinophil activation (assessed by immunohistochemistry). This effect was specific to NGF, since animals given control IgG remained hyperreactive. These data suggest that anticholinergic therapy amplifies eosinophil interactions with airway nerves via NGF. Therefore, therapeutic strategies that target both eosinophil activation and NGF-mediated inflammatory processes in allergic asthma are likely to be beneficial.

  13. Atropine-enhanced, antigen challenge-induced airway hyperreactivity in guinea pigs is mediated by eosinophils and nerve growth factor

    PubMed Central

    Verbout, Norah G.; Jacoby, David B.; Gleich, Gerald J.; Fryer, Allison D.

    2009-01-01

    Although anticholinergic therapy inhibits bronchoconstriction in asthmatic patients and antigen-challenged animals, administration of atropine 1 h before antigen challenge significantly potentiates airway hyperreactivity and eosinophil activation measured 24 h later. This potentiation in airway hyperreactivity is related to increased eosinophil activation and is mediated at the level of the airway nerves. Since eosinophils produce nerve growth factor (NGF), which is known to play a role in antigen-induced airway hyperreactivity, we tested whether NGF mediates atropine-enhanced, antigen challenge-induced hyperreactivity. Antibody to NGF (Ab NGF) was administered to sensitized guinea pigs with and without atropine pretreatment (1 mg/kg iv) 1 h before challenge. At 24 h after challenge, animals were anesthetized, vagotomized, paralyzed, and ventilated. Electrical stimulation of both vagus nerves caused bronchoconstriction that was increased in challenged animals. Atropine pretreatment potentiated antigen challenge-induced hyperreactivity. Ab NGF did not affect eosinophils or inflammatory cells in any group, nor did it prevent hyperreactivity in challenged animals that were not pretreated with atropine. However, Ab NGF did prevent atropine-enhanced, antigen challenge-induced hyperreactivity and eosinophil activation (assessed by immunohistochemistry). This effect was specific to NGF, since animals given control IgG remained hyperreactive. These data suggest that anticholinergic therapy amplifies eosinophil interactions with airway nerves via NGF. Therefore, therapeutic strategies that target both eosinophil activation and NGF-mediated inflammatory processes in allergic asthma are likely to be beneficial. PMID:19447892

  14. Growth-associated protein43 (GAP43) is a biochemical marker for the whole period of fish optic nerve regeneration.

    PubMed

    Kaneda, Manabu; Nagashima, Mikiko; Mawatari, Kazuhiro; Nunome, Tomoya; Muramoto, Kenichiro; Sugitani, Kayo; Kato, Satoru

    2010-01-01

    In adult visual system, goldfish can regrow their axons and fully restore their visual function even after optic nerve transection. The optic nerve regeneration process in goldfish is very long and it takes about a half year to fully recover visual function via synaptic refinement. Therefore, we investigated time course of growth-associated protein 43 (GAP43) expression in the goldfish retina for over 6 months after axotomy. In the control retina, very weak immunoreactivity could be seen in the retinal ganglion cells (RGCs). The immunoreactivity of GAP43 started to increase in the RGCs at 5 days, peaked at 7-20 days and then gradually decreased at 30-40 days after axotomy. The weak but significant immunoreactivity of GAP43 in the RGCs continued during 50-90 days and slowly returned to the control level by 180 days after lesion. The levels of GAP43 mRNA showed a biphasic pattern; a short-peak increase (9-folds) at 1-3 weeks and a long plateau increase (5-folds) at 50-120 days after axotomy. Thereafter, the levels declined to the control value by 180 days after axotomy. The changes of chasing behavior of pair of goldfish with bilaterally axotomized optic nerve also showed a slow biphasic recovery pattern in time course. Although further experiment is needed to elucidate the role of GAP43 in the regrowing axon terminals, the GAP43 is a good biochemical marker for monitoring the whole period of optic nerve regeneration in fish.

  15. Replacement of Nerve-Growth Factor by Ganglionic Non-Neuronal Cells for the Survival In Vitro of Dissociated Ganglionic Neurons

    PubMed Central

    Burnham, Patricia; Raiborn, Charles; Varon, Silvio

    1972-01-01

    Nerve-growth factor is known to cause a considerable increase in the number of neurons putting out processes and surviving in cell cultures of dissociated dorsal-root and sympathetic ganglia from embryonic chicks. Similar effects of nerve-growth factor have now been noted with cultures of dissociated dorsal-root ganglia from newborn mice or rats. In all three sensory ganglionic systems, the effects of the nerve-growth factor on fiber production and neuronal survival could be mimicked, in the absence of the factor, by adequate increase of the non-neuronal cells in the cultures. The results suggest a hypothesis that views the role of the nerve-growth factor as subordinate to that of the non-neuronal cells. PMID:4509314

  16. Upregulation of Nerve Growth Factor in Central Amygdala Increases Sensitivity to Opioid Reward

    PubMed Central

    Bie, Bihua; Wang, Yan; Cai, You-Qing; Zhang, Zhi; Hou, Yuan-Yuan; Pan, Zhizhong Z

    2012-01-01

    The rewarding properties of opioids are essential driving force for compulsive drug-seeking and drug-taking behaviors in the development of opioid-mediated drug addiction. Prior drug use enhances sensitivity to the rewarding effects of subsequently used drugs, increasing vulnerability to relapse. The molecular mechanisms underlying this reward sensitization are still unclear. We report here that morphine that induced reward sensitization, as demonstrated by reinstatement of the behavior of conditioned place preference (CPP) with sub-threshold priming morphine, epigenetically upregulated the output activity of Ngf encoding the nerve growth factor (NGF) by increasing histone H4 acetylation in the rat central nucleus of the amygdala (CeA). NGF locally infused into the CeA mimicked the morphine effect in inducing new functional delta-opioid receptor (DOR) that was required for the reward sensitization, and morphine-induced reward sensitization was inhibited by blocking NGF receptor signaling in the CeA. Histone deacetylase inhibitors that increased the acetylation level at the Ngf promoter and NGF expression in the CeA also induced reward sensitization in a CeA NGF signaling- and DOR-dependent manner. Furthermore, CeA-applied NGF substituted prior morphine to induce reward sensitization in naive rats and also substituted priming morphine to reinstate the CPP induced by prior morphine. Thus, epigenetic upregulation of NGF activity in the CeA may promote the behavior of opioid reward and increase the sensitivity to the rewarding effect of subsequent opioids, a potentially important mechanism in drug addiction. PMID:22871918

  17. Neuroactive conducting scaffolds: nerve growth factor conjugation on active ester-functionalized polypyrrole

    PubMed Central

    Lee, Jae Young; Lee, Joo-Woon; Schmidt, Christine E.

    2009-01-01

    Electrically conductive and biologically active scaffolds are desirable for enhancing adhesion, proliferation and differentiation of a number of cell types such as neurons. Hence, the incorporation of neuroactive molecules into electroconductive polymers via a specific and stable method is essential for neuronal tissue engineering applications. Traditional conjugation approaches dramatically impair conductivities and/or stabilities of the scaffolds and ligands. In this study, we developed copolymers (PPy-NSE) of N-hydroxyl succinimidyl ester pyrrole and regular pyrrole, which can be immobilized with nerve growth factor (NGF) without significantly hindering electroconductivity. The presence of active ester groups was confirmed using reflectance infrared spectroscopy and X-ray photoelectron spectroscopy (XPS) from the copolymers prepared from different monomer compositions. We selected PPy-NSE50 (polymerized from a 50 : 50 monomer ratio of pyrrole : pyrrole-NSE) for further modification with NGF because this copolymer retains good conductivity (approx. 8 S cm−1) and presents active ester groups for NGF immobilization. We tethered NGF on the PPy-NSE50 surface, and found that PC12 cells extended neurites similarly to cells cultured in NGF-containing medium. XPS and enzyme-linked immunosorbent assay confirmed that NGF immobilized via the active ester on the PPy-NSE50 film was stable for up to 5 days in phosphate-buffered saline solution. Also, application of an external electrical potential to NGF-immobilized PPy films did not cause a significant release of NGF nor reduce their neurotrophic activity. This novel scaffold, providing electroconductive and neurotrophic activities, has potential for neural applications, such as tissue engineering scaffolds and biosensors. PMID:19068472

  18. Anti-nerve growth factor antibody attenuates chronic morphine treatment-induced tolerance in the rat.

    PubMed

    Cheppudira, Bopaiah P; Trevino, Alex V; Petz, Lawrence N; Christy, Robert J; Clifford, John L

    2016-09-05

    Nerve growth factor (NGF) is known to induce inflammation and pain; however its role in opioid-induced tolerance has not been studied. This study investigated the effects of an anti-NGF neutralizing antibody on the development of tolerance following chronic morphine treatment in naïve rats. Four groups of rats were used in this study; one treated with saline alone, one with 10 mg/kg of morphine, one with 10 μg of anti-NGF and the other with 10 mg/kg of morphine + 10 μg of anti-NGF, twice per day for 5 days. The route of treatment was subcutaneous (S.C.) for morphine and saline, and intraperitoneal (i.p.) for anti-NGF. Response to a noxious thermal stimulus during the course of drug treatment was assessed (Hargreaves' test). Further, the change in the NGF levels in the lumbar spinal cord was measured by ELISA. Our results showed that repeated administration of morphine produced an apparent tolerance which was significantly attenuated by co-administration of anti-NGF (P < 0.001). Additionally, the area under the curve (AUC) of the analgesic effect produced by the combination of morphine and anti-NGF was significantly (P < 0.001) greater than for saline controls and chronic morphine treated rats. Moreover, the level of NGF in the spinal cord of chronic morphine treated rats was significantly higher (P < 0.05) than in both the saline control group and the group receiving simultaneous administration of anti-NGF with morphine. These results indicate that anti-NGF has the potential to attenuate morphine-induced tolerance behavior by attenuating the effects of NGF at the spinal level. Taken together, our study strongly suggests that the NGF signaling system is a potential novel target for treating opioid-induced tolerance.

  19. Persistent Nociception Triggered by Nerve Growth Factor (NGF) Is Mediated by TRPV1 and Oxidative Mechanisms.

    PubMed

    Eskander, Michael A; Ruparel, Shivani; Green, Dustin P; Chen, Paul B; Por, Elaine D; Jeske, Nathaniel A; Gao, Xiaoli; Flores, Eric R; Hargreaves, Kenneth M

    2015-06-03

    Nerve growth factor (NGF) is elevated in certain chronic pain conditions and is a sufficient stimulus to cause lasting pain in humans, but the actual mechanisms underlying the persistent effects of NGF remain incompletely understood. We developed a rat model of NGF-induced persistent thermal hyperalgesia and mechanical allodynia to determine the role of transient receptor potential vanilloid 1 (TRPV1) and oxidative mechanisms in the persistent effects of NGF. Persistent thermal hypersensitivity and mechanical allodynia require de novo protein translation and are mediated by TRPV1 and oxidative mechanisms. By comparing effects after systemic (subcutaneous), spinal (intrathecal) or hindpaw (intraplantar) injections of test compounds, we determined that TRPV1 and oxidation mediate persistent thermal hypersensitivity via peripheral and spinal sites of action and mechanical allodynia via only a spinal site of action. Therefore, NGF-evoked thermal and mechanical allodynia are mediated by spatially distinct mechanisms. NGF treatment evoked sustained increases in peripheral and central TRPV1 activity, as demonstrated by increased capsaicin-evoked nocifensive responses, increased calcitonin gene-related peptide release from hindpaw skin biopsies, and increased capsaicin-evoked inward current and membrane expression of TRPV1 protein in dorsal root ganglia neurons. Finally, we showed that NGF treatment increased concentrations of linoleic and arachidonic-acid-derived oxidized TRPV1 agonists in spinal cord and skin biopsies. Furthermore, increases in oxidized TRPV1-active lipids were reduced by peripheral and spinal injections of compounds that completely blocked persistent nociception. Collectively, these data indicate that NGF evokes a persistent nociceptive state mediated by increased TRPV1 activity and oxidative mechanisms, including increased production of oxidized lipid TRPV1 agonists.

  20. Internalization of nerve growth factor by pheochromocytoma PC12 cells: absence of transfer to the nucleus.

    PubMed

    Rohrer, H; Schäfer, T; Korsching, S; Thoenen, H

    1982-06-01

    The intracellular distribution of 125I-labeled nerve growth factor (NGF) in rat pheochromocytoma PC12 cells was studied by quantitative electron microscopic (EM) autoradiography and by subcellular fractionation. PC12 cells were grown as monolayer cultures in medium supplemented with serum in the presence of 125I-NGF. EM autoradiography showed that 125I-NGF was localized at the plasma membrane and cytoplasmic compartments but did not accumulate in the nuclear chromatin or in the nuclear membrane compartment of cells analyzed after 1 hr and 1, 2, and 8 d of incubation with 125I-NGF. 125I-NGF also was not detected in nuclear subcellular fractions prepared from cells grown in serum-supplemented medium either in suspension for 1 d or in monolayer cultures for 1 to 8 d. In contrast, and in confirmation of the results of Yankner and Shooter (Yankner, B. A., and E. M. Shooter (1979) Pro. Natl. Acad. Sci. U. S. A. 76: 1269-1273), about 60% of the cell-bound 125I-NGF was found in the nuclear pellet after cell fractionation if the cells had been kept previously in suspension for 1 d in phosphate-buffered saline supplemented with 0.2% glucose, 0.1% bovine serum albumin, and 125I-NGF. The ultrastructure of PC12 cells grown under such conditions, however, revealed signs of varying degrees of damage. Autoradiography of the nuclear pellet from these cells showed the grains to be located mainly over damaged nuclei or over cell debris between nuclei. It is concluded that NGF, after binding to specific receptors at the plasma membrane, is transferred to membrane-confined cytoplasmic compartments but does not have to be transferred further to the nuclear membrane or to the nuclear chromatin as a prerequisite for its physiological action.

  1. Radix Bupleuri ameliorates depression by increasing nerve growth factor and brain-derived neurotrophic factor

    PubMed Central

    Wang, Xia; Feng, Qing; Xiao, Yong; Li, Ping

    2015-01-01

    Background: Chinese herb Radix Bupleuri has been regarded effective to improve treatment of depression, but the molecular mechanism remains unknown. Low levels of nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) increase the likelihood of developing the depression. Therefore, we want to know whether Radix Bupleuri affects the levels of these factors. Methods: A total 160 hemodialysis patients were diagnosed with depression and randomly assigned to two groups: Radix Bupleuri group (received 1 g root power of Radix Bupleuri in a capsule daily Radix Bupleuri) and control group (receive placebo). Results: After three-month follow-up, the patients who received Radix Bupleuri had greater improvement in depression symptoms, anxiety symptoms and general functioning via controls after three-month follow-up (P < 0.05). Serum NGF levels were significantly higher in subjects accepted Radix Bupleuris (178.64 ± 52.18 pg/mL) when compared to a control (103.54 ± 31.23 pg/ml) (P < 0.01). Similarly, serum BDNF levels were significantly higher in subjects accepted Radix Bupleuris (1635.26 ± 121.66 pg/ml) when compared to a control (516.38 ± 44.89 pg/ml) (P < 0.01). The serum levels of NGF and BDNF were negatively related with Montgomery-Asberg Depression Rating Scale (MADRS) and positively related with scores of RAND-36 item Health Survey (RAND-36) (P < 0.01). Conclusion: Thus, Radix Bupleuri ameliorates the patients with depression by increasing serum levels of NGF and BDNF. Radix Bupleuri should be developed a new drug for the therapy of depression. PMID:26309578

  2. Urinary nerve growth factor levels in overactive bladder syndrome and lower urinary tract disorders.

    PubMed

    Liu, Hsin-Tzu; Chen, Chia-Yen; Kuo, Hann-Chorng

    2010-12-01

    Overactive bladder (OAB) is a syndrome based on self-reported symptoms of urgency and frequency with or without urge incontinence. Although urgency is the core symptom of OAB, patients might have difficulty to distinguish urgency from the urge to void. Urodynamic study is a useful diagnostic tool to discover detrusor overactivity (DO) in patients with OAB; however, not all OAB patients have DO. Therefore, a more objective and non-invasive way to diagnose and assess OAB including DO is needed. Recent research has focused on urinary biomarkers in assessment of OAB. Urinary nerve growth factor (NGF) level increases in patients with OAB-wet, bladder outlet obstruction, mixed urinary incontinence and urodynamic DO. Urinary NGF levels are correlated with severity of OAB symptoms. In patients with OAB and DO who have been well treated with antimuscarinics or botulinum toxin injection, urinary NGF levels have been shown to decrease significantly in association with reduction of urgency severity. However, not all patients with OAB have an elevated urinary NGF level. It might also be increased in patients with interstitial cystitis/painful bladder syndrome, cerebrovascular accident and lower urinary tract diseases such as urinary tract stone, bacterial infection and urothelial tumor. It is possible to use urinary NGF levels as a bio-marker for diagnosis of OAB as well as for the assessment of therapeutic outcome in patients with OAB or DO. Here, we review the latest medical advances in this field. Copyright © 2010 Formosan Medical Association & Elsevier. Published by Elsevier B.V. All rights reserved.

  3. Neurosteroid dehydroepiandrosterone interacts with nerve growth factor (NGF) receptors, preventing neuronal apoptosis.

    PubMed

    Lazaridis, Iakovos; Charalampopoulos, Ioannis; Alexaki, Vassilia-Ismini; Avlonitis, Nicolaos; Pediaditakis, Iosif; Efstathopoulos, Paschalis; Calogeropoulou, Theodora; Castanas, Elias; Gravanis, Achille

    2011-04-01

    The neurosteroid dehydroepiandrosterone (DHEA), produced by neurons and glia, affects multiple processes in the brain, including neuronal survival and neurogenesis during development and in aging. We provide evidence that DHEA interacts with pro-survival TrkA and pro-death p75(NTR) membrane receptors of neurotrophin nerve growth factor (NGF), acting as a neurotrophic factor: (1) the anti-apoptotic effects of DHEA were reversed by siRNA against TrkA or by a specific TrkA inhibitor; (2) [(3)H]-DHEA binding assays showed that it bound to membranes isolated from HEK293 cells transfected with the cDNAs of TrkA and p75(NTR) receptors (K(D): 7.4 ± 1.75 nM and 5.6 ± 0.55 nM, respectively); (3) immobilized DHEA pulled down recombinant and naturally expressed TrkA and p75(NTR) receptors; (4) DHEA induced TrkA phosphorylation and NGF receptor-mediated signaling; Shc, Akt, and ERK1/2 kinases down-stream to TrkA receptors and TRAF6, RIP2, and RhoGDI interactors of p75(NTR) receptors; and (5) DHEA rescued from apoptosis TrkA receptor positive sensory neurons of dorsal root ganglia in NGF null embryos and compensated NGF in rescuing from apoptosis NGF receptor positive sympathetic neurons of embryonic superior cervical ganglia. Phylogenetic findings on the evolution of neurotrophins, their receptors, and CYP17, the enzyme responsible for DHEA biosynthesis, combined with our data support the hypothesis that DHEA served as a phylogenetically ancient neurotrophic factor.

  4. Nerve growth factor attenuates cholinergic deficits following traumatic brain injury in rats.

    PubMed

    Dixon, C E; Flinn, P; Bao, J; Venya, R; Hayes, R L

    1997-08-01

    Traumatic brain injury (TBI) results in chronic derangements in central cholinergic neurotransmission that may contribute to posttraumatic memory deficits. Intraventricular cannula (IVC) nerve growth factor (NGF) infusion can reduce axotomy-induced spatial memory deficits and morphologic changes observed in medial septal cholinergic neurons immunostained for choline acetyltransferase (ChAT). We examined the efficacy of NGF to (1) ameliorate reduced posttraumatic spatial memory performance, (2) release of hippocampal acetylcholine (ACh), and (3) ChAT immunoreactivity in the rat medial septum. Rats (n = 36) were trained prior to TBI on the functional tasks and retested on Days 1-5 (motor) and on Day 7 (memory retention). Immediately following injury, an IVC and osmotic pump were implanted, and NGF or vehicle was infused for 7 days. While there were no differences in motor performance, the NGF-treated group had significantly better spatial memory retention (P < 0.05) than the vehicle-treated group. The IVC cannula was then removed on Day 7, and a microdialysis probe was placed into the dorsal hippocampus. After a 22-h equilibration period, samples were collected prior to and after administration of scopolamine (1 mg/kg), which evoked ACh release by blocking autoreceptors. The posttraumatic reduction in scopolamine-evoked ACh release was completely reversed with NGF. Injury produced a bilateral reduction in the number and cross-sectional area of ChAT immunopositive medial septal neurons that was reversed by NGF treatment. These data suggest that cognitive but not motor deficits following TBI are, in part, mediated by chronic deficits in cholinergic systems that can be modulated by neurotrophic factors such as NGF.

  5. Serum nerve growth factor levels in autistic children in Turkish population: A preliminary study

    PubMed Central

    Dinçel, Nida; Ünalp, Aycan; Kutlu, Ayşe; Öztürk, Aysel; Uran, Nedret; Ulusoy, Sadık

    2013-01-01

    Background & objectives: It has been hypothesized that abnormal levels of serum nerve growth factor (NGF) may represent a serological marker for autistic children who may develop cognitive impairment, regression and finally epilepsy. The objective of this preliminary study was to measure serum NGF concentrations of autistic children and compare these levels with those of healthy children. Methods: Consecutive children who were referred to the Paediatric Neurology and Child Psychiatry Policlinics of Dr. Behçet Uz Child Disease and Pediatric Surgery Training and Research Hospital, Turkey between February and September 2008 were included in the study. Serum samples were analyzed for NGF levels using ChemiKine NGF Sandwich ELISA Kit. Comparisons between the study and the control groups were made using student's t test and Chi-square test. Results: Forty-nine autistic children and an equal number of healthy children (control group) were included in the study. No significant difference was found between the study and the control groups in terms of children's age, while number of boys was significantly higher (P<0.05) in the study group. Average serum NGF concentrations were 46.94 ± 51.40 and 32.94 ± 12.48 pg/ml in the study and control group, respectively. Serum NGF concentrations were significantly higher (P<0.05) in the study group compared with the control group. Interpretation & conclusions: Our preliminary findings show that enhanced serum NGF concentration may be used as a potential diagnostic tool in autism, however, further studies including a large number of patients are required to confirm the findings. PMID:24521633

  6. Intrathecal lidocaine neurotoxicity: combination with bupivacaine and ropivacaine and effect of nerve growth factor.

    PubMed

    Zhao, Guangyi; Ding, Xudong; Guo, Yao; Chen, Weimin

    2014-09-01

    The study aims to investigate the neurotoxicity induced by combined use of intrathecal lidocaine with bupivacaine and ropivacaine, and to examine the effect of nerve growth factor (NGF) on lidocaine-induced neurotoxicity. All rats received intrathecal infusion of anesthetics and NGF. To study the neurotoxicity of combined use of lidocaine with bupivacaine and ropivacaine, rats received saline, 5% lidocaine, 1.065% bupivacaine, 1.5% ropivacaine, 5% lidocaine+bupivacaine, or 5% lidocaine+1.5% ropivacaine. To study the neurotoxicity of different proportions of lidocaine and bupivacaine, mixtures were made by mixing 10% lidocaine and 2.5% bupivacaine in ratios of 1:3, 1:2, 1:1, 2:1 and 3:1 by volume. To study the effect of NGF on lidocaine-induced neurotoxicity, rats received saline or 10 μg NGF for 1, 2, 5 and 8 days. The neurotoxicity of lidocaine was significantly increased when combined with ropivacaine. A mixture of lidocaine and bupivacaine in a ratio of ≤1:1 did not significantly increase lidocaine-induced neurotoxicity. NGF significantly reduced lidocaine-induced neurobehavioral and morphological damage in the spinal cord. This was accompanied by downregulation of caspase 3 expression. Ropivacaine is not safe when intrathecally administered with lidocaine at the concentrations used in this study. Bupivacaine may be safely used with lidocaine at a ratio of 1:1. NGF can reduce lidocaine-induced neurotoxicity, possibly via inhibition of caspase 3-mediated apoptosis. Copyright © 2014 Elsevier Inc. All rights reserved.

  7. Single cycle structure-based humanization of an anti-nerve growth factor therapeutic antibody.

    PubMed

    Covaceuszach, Sonia; Marinelli, Sara; Krastanova, Ivet; Ugolini, Gabriele; Pavone, Flaminia; Lamba, Doriano; Cattaneo, Antonino

    2012-01-01

    Most forms of chronic pain are inadequately treated by present therapeutic options. Compelling evidence has accumulated, demonstrating that Nerve Growth Factor (NGF) is a key modulator of inflammatory and nociceptive responses, and is a promising target for the treatment of human pathologies linked to chronic and inflammatory pain. There is therefore a growing interest in the development of therapeutic molecules antagonising the NGF pathway and its nociceptor sensitization actions, among which function-blocking anti-NGF antibodies are particularly relevant candidates.In this respect, the rat anti-NGF αD11 monoclonal antibody (mAb) is a potent antagonist, able to effectively antagonize rodent and human NGF in a variety of in vitro and in vivo systems. Here we show that mAb αD11 displays a significant analgesic effect in two different models of persistent pain in mice, with a remarkable long-lasting activity. In order to advance αD11 mAb towards its clinical application in man, anti-NGF αD11 mAb was humanized by applying a novel single cycle strategy based on the a priori experimental determination of the crystal and molecular structure of the parental Fragment antigen-binding (Fab). The humanized antibody (hum-αD11) was tested in vitro and in vivo, showing that the binding mode and the NGF neutralizing biological activities of the parental antibody are fully preserved, with even a significant affinity improvement. The results firmly establish hum-αD11 as a lead candidate for clinical applications in a therapeutic area with a severe unmet medical need. More generally, the single-cycle structure-based humanization method represents a considerable improvement over the standard humanization methods, which are intrinsically empirical and require several refinement cycles.

  8. Noninflammatory upregulation of nerve growth factor underlies gastric hypersensitivity induced by neonatal colon inflammation

    PubMed Central

    Li, Qingjie; Winston, John H.

    2015-01-01

    Gastric hypersensitivity is one of the key contributors to the postprandial symptoms of epigastric pain/discomfort, satiety, and fullness in functional dyspepsia patients. Epidemiological studies found that adverse early-life experiences are risk factors for the development of gastric hypersensitivity. Preclinical studies found that neonatal colon inflammation elevates plasma norepinephrine (NE), which upregulates expression of nerve growth factor (NGF) in the muscularis externa of the gastric fundus. Our goal was to investigate the cellular mechanisms by which NE upregulates the expression of NGF in gastric hypersensitive (GHS) rats, which were subjected previously to neonatal colon inflammation. Neonatal colon inflammation upregulated NGF protein, but not mRNA, in the gastric fundus of GHS rats. Western blotting showed upregulation of p110γ of phosphatidylinositol 4,5-bisphosphate 3-kinase (PI3K), phosphoinositide-dependent kinase-1 (PDK1), pAKT(Ser473), and phosphorylated 4E-binding protein (p4E-BP1)(Thr70), suggesting AKT activation and enhanced NGF protein translation. AKT inhibitor MK-2206 blocked the upregulation of NGF in the fundus of GHS rats. Matrix metalloproteinase 9 (MMP-9), the major NGF-degrading protease, was suppressed, indicating that NGF degradation was impeded. Incubation of fundus muscularis externa with NE upregulated NGF by modulating the protein translation and degradation pathways. Yohimbine, an α2-adrenergic receptor antagonist, upregulated plasma NE and NGF expression by activating the protein translation and degradation pathways in naive rats. In contrast, a cocktail of adrenergic receptor antagonists suppressed the upregulation of NGF by blocking the activation of the protein translation and degradation pathways. Our findings provide evidence that the elevation of plasma NE induces NGF expression in the gastric fundus. PMID:26608656

  9. Nerve growth factor translates stress response and subsequent murine abortion via adhesion molecule-dependent pathways.

    PubMed

    Tometten, Mareike; Blois, Sandra; Kuhlmei, Arne; Stretz, Anna; Klapp, Burghard F; Arck, Petra C

    2006-04-01

    Spontaneous abortion is a frequent threat affecting 10%-25% of human pregnancies. Psychosocial stress has been suggested to be attributable for pregnancy losses by challenging the equilibrium of systems mandatory for pregnancy maintenance, including the nervous, endocrine, and immune system. Strong evidence indicates that stress-triggered abortion is mediated by adhesion molecules, i.e., intercellular adhesion molecule 1 (ICAM1) and leukocyte function associated molecule 1, now being referred to as integrin alpha L (ITGAL), which facilitate recruitment of inflammatory cells to the feto-maternal interface. The neurotrophin beta-nerve growth factor (NGFB), which has been shown to be upregulated in response to stress in multiple experimental settings including in the uterine lining (decidua) during pregnancy, increases ICAM1 expression on endothelial cells. Here, we investigated whether and how NGFB neutralization has a preventive effect on stress-triggered abortion in the murine CBA/J x DBA/2J model. We provide experimental evidence that stress exposure upregulates the frequency of abortion and the expression of uterine NGFB. Further, adhesion molecules ICAM1 and selectin platelet (SELP, formerly P-Selectin) and their ligands ITGAL and SELP ligand (SELPL, formerly P selectin glycoprotein ligand 1) respectively increase in murine deciduas in response to stress. Subsequently, decidual cytokines are biased toward a proinflammatory and abortogenic cytokine profile. Additionally, a decrease of pregnancy protective CD8alpha(+) decidual cells is present. Strikingly, all such uterine stress responses are abrogated by NGFB neutralization. Hence, NGFB acts as a proximal mediator in the hierarchical network of immune rejection by mediating an abortogenic environment comprised of classical signs of neurogenic inflammation.

  10. Nerve Growth Factor Promotes Reorganization of the Axonal Microtubule Array at Sites of Axon Collateral Branching

    PubMed Central

    Ketschek, Andrea; Jones, Steven; Spillane, Mirela; Korobova, Farida; Svitkina, Tatyana; Gallo, Gianluca

    2015-01-01

    The localized debundling of the axonal microtubule array and the entry of microtubules into axonal filopodia are two defining features of collateral branching. We report that nerve growth factor (NGF), a branch inducing signal, increases the frequency of microtubule debundling along the axon shaft of chicken embryonic sensory neurons. Sites of debundling correlate strongly with the localized targeting of microtubules into filopodia. Platinum replica electron microscopy suggests physical interactions between debundled microtubules and axonal actin filaments. However, as evidenced by depolymerization of actin filaments and inhibition of myosin II, actomyosin force generation does not promote debundling. In contrast, loss of actin filaments or inhibition of myosin II activity promotes debundling, indicating that axonal actomyosin forces suppress debundling. MAP1B is a microtubule associated protein that represses axon branching. Following treatment with NGF, microtubules penetrating filopodia during the early stages of branching exhibited lower levels of associated MAP1B. NGF increased and decreased the levels of MAP1B phosphorylated at a GSK-3β site (pMAP1B) along the axon shaft and within axonal filopodia, respectively. The levels of MAP1B and pMAP1B were not altered at sites of debundling, relative to the rest of the axon. Unlike the previously determined effects of NGF on the axonal actin cytoskeleton, the effects of NGF on microtubule debundling were not affected by inhibition of protein synthesis. Collectively, these data indicate that NGF promotes localized axonal microtubule debundling, that actomyosin forces antagonize microtubule debundling and that NGF regulates pMAP1B in axonal filopodia during the early stages of collateral branch formation. PMID:25846486

  11. Can proline-rich polypeptide complex mimic the effect of nerve growth factor?

    PubMed

    Zabłocka, Agnieszka; Urbaniak, Anna; Kuropatwa, Marianna; Zyzak, Joanna; Rossowska, Joanna; Janusz, Maria

    2014-01-01

    Naturally occurring compounds that can act as prosurvival factors and neurite formation stimulants in the conditions of reduced neurotrophins production are important both in neuronal protection and therapy of neurodegenerative disorders. Therefore, the role of proline-rich polypeptide complex (PRP) and its nonapeptide fragment (NP) in the promotion of pheochromocytoma cell line (PC12) survival and neurite outgrowth pathway is presented. It was shown that PRP/NP did not affect the neuronal nitric oxide synthase (nNOS) at the transcriptional and protein level. However, the activity of nNOS and intracellular nitric oxide (NO) concentration was markedly increased after treatment of PC12 cells with peptides. This reaction was inhibited by L-NAME-nNOS inhibitor. It was shown that PRP and NP induce the soluble guanylyl cyclase to release higher amount of cyclic GMP (cGMP), and subsequently, the increased phosphorylation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) is observed. This effect was abolished by both U0126 (inhibitor of ERK1/2) and also by L-NAME. Reduction of ERK1/2 activity observed in the presence of nNOS inhibitor suggests that its activation is NO-dependent. The presented results shed some light on the mechanism of action of PRP complex. PRP and NP can activate NO/cGMP/ERK1/2 signaling pathway, similarly to nerve growth factor (NGF). The prosurvival action and short fibers formation suggest the role of PRP and NP in neuroprotection and the initiation of neuritogenesis. They can also participate in the amplification of signals controlling the survival and differentiation of neurons effect when the deficit of NGF takes place.

  12. Exploring the role of nerve growth factor in multiple sclerosis: implications in myelin repair.

    PubMed

    Acosta, C M R; Cortes, C; MacPhee, H; Namaka, M P

    2013-12-01

    Multiple sclerosis (MS) is a chronic disease resulting from targeted destruction of central nervous system (CNS) myelin. MS is suggested to be an autoimmune disease involving the pathogenic activation of CD4(+) T cells by a foreign antigen in the peripheral blood. The activated CD4(+) T cells liberate inflammatory cytokines that facilitate the breakdown of the blood-brain barrier (BBB) promoting their passage into the CNS. Inside the CNS, CD4(+) T cells become re-activated by myelin proteins sharing a similar structure to the foreign antigen that initially triggered the immune response. The CD4(+) T cells continue to liberate inflammatory cytokines, such as tumor necrosis factor α (TNFα), which activates macrophages and antibodies responsible for the phagocytosis of myelin. Acute CNS lesions can be re-myelinated, however, the repair of chronic demyelinating lesions is limited, leading to permanent neurological deficits. Although current MS treatments reduce severity and slow disease progression, they do not directly repair damaged myelin. Henceforth, recent treatment strategies have focused on neurotrophins, such as nerve growth factor (NGF) for myelin repair. NGF promotes axonal regeneration, survival, protection and differentiation of oligodendrocytes (OGs) and facilitates migration and proliferation of oligodendrocyte precursors (OPs) to the sites of myelin damage. NGF also directly regulates key structural proteins that comprise myelin. Interestingly, NGF also induces the production of brain-derived neurotrophic factor (BDNF), another integral neurotrophin involved in myelination. The intricate signaling between neurotrophins and cytokines that governs myelin repair supports the role of NGF as a leading therapeutic candidate in white matter disorders, such as MS.

  13. Discoordinate regulation of different K channels in cultured rat skeletal muscle by nerve growth factor.

    PubMed

    Vigdor-Alboim, S; Rothman, C; Braiman, L; Bak, A; Langzam, L; Yosef, O; Sterengarz, B B; Nawrath, H; Brodie, C; Sampson, S R

    1999-05-01

    We investigated the effects of nerve growth factor (NGF) on expression of K+ channels in cultured skeletal muscle. The channels studied were (1) charybdotoxin (ChTx)-sensitive channels by using a polyclonal antibody raised in rabbits against ChTx, (2) Kv1.5 voltage-sensitive channels, and (3) apamin-sensitive (afterhyperpolarization) channels. Crude homogenates were prepared from cultures made from limb muscles of 1-2-day-old rat pups for identification of ChTx-sensitive and Kv1.5 channels by Western blotting techniques. Apamin-sensitive K+ channels were studied by measurement of specific [125I]-apamin binding by whole cell preparations. ChTx-sensitive channels display a fusion-related increase in expression, and NGF downregulates these channels in both myoblasts and myotubes. Voltage-dependent Kv1.5 channel expression is low in myoblasts and increases dramatically with fusion; NGF induces early expression of these channels and causes expression after fusion to increase even further. NGF downregulates apamin-sensitive channels. NGF increases the rate of fall of the action potential recorded intracellularly from single myotubes with intracellular microelectrodes. The results confirm and extend those of previous studies in showing a functional role for NGF in the regulation of membrane properties of skeletal muscle. Moreover, the findings demonstrate that the different K+ channels in this preparation are regulated in a discoordinate manner. The divergent effects of NGF on expression of different K+ channels, however, do not appear sufficient to explain the NGF-induced increase in the rate of fall of the action potential. The changes during the falling phase may rather be due to increases in channel properties or may result from an increased driving force on the membrane potential secondary to the NGF-induced hyperpolarization.

  14. Maternal nerve growth factor levels during pregnancy in women with preeclampsia: A longitudinal study.

    PubMed

    D'souza, Vandita; Kilari, Anitha; Pisal, Hemlata; Patil, Vidya; Mehendale, Savita; Wagh, Girija; Gupte, Sanjay; Joshi, Sadhana

    2015-12-01

    Preeclampsia (PE) is characterized by hypertension and proteinuria. Improper development of the placenta due to altered angiogenesis is the main culprit in PE. Nerve growth factor (NGF) is an angiogenic factor which is expressed and localized in the placenta. Our earlier cross sectional study has shown altered NGF levels at delivery in women with PE. However, there are no studies on NGF levels in PE early in pregnancy before manifestation of the disease. Thus, there is a need to examine the role of NGF in vascular development during different stages of gestation in PE. A longitudinal study was carried out where pregnant women were enrolled from two major hospitals from Pune, Bharati hospital and Gupte hospital. They were followed at three different time points [16-20 weeks (T1), 26-30 weeks (T2) and at delivery (T3)] during pregnancy and maternal blood at every time point and cord blood at delivery was collected and processed. This study included normotensive women (n=88) and women with PE (n=48). NGF levels were measured from maternal and cord plasma using the Emax Immuno Assay System (Promega). The data was analyzed using the SPSS/PC+ package (Version 20.0, Chicago, IL, USA). Maternal NGF levels did not change at all time points while cord NGF levels were higher (p<0.05) in women with PE. Further, maternal NGF levels were negatively associated with blood pressure while cord NGF levels were positively associated with baby head circumference. Our data suggests that there may possibly be a compensatory role for NGF in the foeto-placental circulation in PE.

  15. Concentration-dependent effect of nerve growth factor on cell fate determination of neural progenitors.

    PubMed

    Zhang, Lei; Jiang, Hui; Hu, Zhengqing

    2011-10-01

    Stem cell-based spiral ganglion neuron (SGN) replacement therapy has been proposed to be a promising strategy to restore hearing either via replacing degenerated neurons or by improving the efficacy of cochlear implants which rely on functional neurons. However, lack of suitable donor cells and low survival rate of implanted cells are the major obstacles to successful implementation of therapeutic transplantation. The present study investigated the potential of mouse inner ear statoacoustic ganglion (SAG)-derived neural progenitors (NPs) to differentiate toward SGN-like glutamatergic cells and the influence to cell survival and differentiation when nerve growth factor (NGF) was supplied. We found that SAG-NPs could form neurospheres, proliferate, and differentiate into cells expressing neuronal protein neurofilament and β-III tubulin. NGF affected the cell fate of SAG-NP in a concentration-dependent manner in vitro. Low concentration of NGF (2-5 ng/mL) promoted cell proliferation. Medium concentration of NGF (20-40 ng/mL) stimulated cells to differentiate into bi-polar SGN-like cells expressing glutamatergic proteins. High concentration of NGF (100 ng/mL) could rescue cells from induced apoptosis. In the in vivo study, NGF (100 ng/mL) dramatically enhanced SAG-NP survival rate after implantation into adult mammalian inner ear. This finding raises the possibility to further induce these NPs to differentiate into SGN-like neurons in future in vivo study. In conclusion, given the capability of proliferation and differentiation into SGN-like cells with the supplement of NGF in vitro, SAG-NPs can serve as donor cells in stem cell-based SGN replacement therapy. NGF improved the survival of SAG-NPs not only in vitro but also in vivo.

  16. Microtubules and Microfilaments in Fixed and Permeabilized Cells are Selectively Decorated by Nerve Growth Factor

    NASA Astrophysics Data System (ADS)

    Nasi, S.; Cirillo, D.; Naldini, L.; Marchisio, P. C.; Calissano, P.

    1982-02-01

    A specific antibody against nerve growth factor (NGF) and indirect immunofluorescence microscopy have been used to follow the in vitro binding of NGF to cells made permeable to large molecules. All cells tested, both target (sensory neurons and PC12 cells) and nontarget (3T3, BKH 2I, C6 glioma cells), revealed a decoration of cytoskeletal structures which on the basis of their form, reactivity with antibodies, and sensitivity to specific drugs may be identified as microtubules (MTs) and microfilaments (MFs). The decoration of either structure depends on the fixation and permeabilization conditions: MFs, in the form of stress fibers, are stained by NGF when the plasma membrane is permeabilized with methanol/acetone; MTs become intensely stained when the plasma membrane is solubilized with a nonionic detergent in the presence of a MT-stabilizing medium. The two procedures do not affect the staining of these structures with specific antibodies. Binding of 125I-labeled NGF to PC12 cells was not competitively inhibited by a 100-fold excess of several positively charged proteins but it was markedly decreased in the presence of DNase I. 125I-Labeled NGF interacted with MTs and F-actin (fixed with paraformaldehyde) in a range of concentrations similar to that used for their cellular localization with NGF-anti-NGF. Our studies show that the specificity and affinity of NGF binding to MTs and MFs is in the range of that of antibodies against tubulin and actin. The possible relevance of these findings to the mechanism of action of NGF in target cells is discussed.

  17. [Clinical efficacy of mouse nerve growth factor in the treatment of sudden deafness].

    PubMed

    Xu, Ting; Xiao, Dajiang; Wu, Sihai; Yuan, Yuan

    2014-05-01

    To study the clinical efficacy of mouse nerve growth factor (NGF) in the treatment of sudden deafness. A retrospective analysis was performed on 115 cases of hospitalized patients who were suffered from sudden deafness. Patients were divided into two groups according to treatment medicine. Control group: patients were treated with intravenous vasodilators, energy mixture, steroid pulse therapy, and methylcobalamin neurotrophic therapy. NGF group: intramuscular NGF treatment was added on the basis of conventional therapy mentioned above. Both treatments lasted 14 days, the total efficiency were compared. Patients were further divided into sub-groups according to age, duration and the level of pre-treatment PTA, and the treatment efficiency was further compared. By SPSS 11.0 statistical analysis, a P < 0.05 was considered as statistical significant difference. (1) The total efficiency of NGF group was significantly higher than control group. (2) Regard of age, the efficiency of NGF treatment group was significantly higher than control group. (3) For the patients whose duration were less than 7 d, or the PTA < or = 60 dBHL, the efficiency of NGF group were significantly higher. For the patients whose duration were more than 7 d, or the PTA>60 dBHL, the efficiency of NGF therapy was not superior to the traditional treatment. NGF can significantly improve the symptom of patients with short duration or low PTA. For this kind of patients, NGF adjuvant therapy should be recommended. For the patients with longer duration and higher level of PTA, NGF therapy is not advocated. NGF treatment should not be in consideration of the age.

  18. Lower Levels of Urinary Nerve Growth Factor Might Predict Recurrent Urinary Tract Infections in Women

    PubMed Central

    2016-01-01

    Purpose: To investigate the changes in urinary nerve growth factor (uNGF) levels after acute urinary tract infection (UTI) and to assess the role of uNGF in predicting UTI recurrence in women. Methods: Women with uncomplicated, symptomatic UTIs were enrolled. Cephalexin 500 mg (every 6 hours) was administered for 7–14 days to treat acute UTIs. Subsequently, the patients were randomized to receive either sulfamethoxazole/trimethoprim 800 mg/160 mg daily at bedtime, or celecoxib 200 mg daily for 3 months and were monitored for up to 12 months. NGF levels in the urine were determined at baseline, 1, 4, and 12 weeks after the initiation of prophylactic therapy, and were compared between women with first-time UTIs and recurrent UTIs, sulfamethoxazole/trimethoprim and celecoxib-treated women, and no UTI recurrence and UTI recurrence that occurred during the follow-up period. Twenty women free of UTIs served as controls. Results: A total of 139 women with UTI and 20 controls were enrolled in the study, which included 50 women with a first-time UTI and 89 women with recurrent UTIs. Thirty-seven women completed the study. Women with recurrent UTIs (n=23) had a trend of lower uNGF levels than women with first-time UTIs (n=14). During follow-up, 9 women had UTI recurrence. The serial uNGF levels in women with UTI recurrence were significantly lower than those in women who did not have UTI recurrence during the follow-up period. Conclusions: The lower levels of uNGF in women with recurrent UTI and the incidence of UTI recurrence during follow-up suggest that lower uNGF might reflect the defective innate immunity in women with recurrent UTI. PMID:27032555

  19. Salivary nerve growth factor response to intense stress: effect of sex and body mass index.

    PubMed

    Taylor, Marcus K; Laurent, Heidemarie K; Larson, Gerald E; Rauh, Mitchell J; Hiller Lauby, Melissa D; Granger, Douglas A

    2014-05-01

    Ample evidence links stress to psychiatric and neurological disease. Although many studies examine stress hormone secretion and receptor activity, exciting new developments signify a shift in focus to neuromodulatory systems influencing neuronal development, survival, and neuroplasticity. The purpose of this study was to characterize salivary nerve growth factor (sNGF) responses to intense stress exposure in healthy military members undergoing survival training. A second purpose was to explore effects of age, sex, education, and body mass index (BMI). One hundred sixteen military members (80% male) were studied before, during, and 24 h after a stressful mock-captivity exercise. sNGF was measured at all three time points. Reactivity, recovery, and residual elevation of sNGF were computed. General linear modeling with repeated measures evaluated effect of stress exposure, as well as the roles of age, sex, education, and BMI. sNGF increased 137% from baseline to intense stress. During recovery, sNGF remained elevated an average of 67% above baseline (i.e., residual elevation). Men showed greater sNGF reactivity than women quantified by larger absolute T1-T2Δ (+148.1 pg/mL vs. +64.9 pg/mL, p<0.017). A noteworthy trend of higher sNGF concentrations in low BMI participants was observed (p=0.058). No effects of age or education were shown. This study shows substantial reactivity and residual elevation of sNGF in response to intense stress exposure in healthy humans. Further research is needed to refine the sNGF assay, fully characterize the sNGF stress response, delineate correlates and mechanisms, and validate therapeutic applications.

  20. Nerve growth factor regulates axial rotation during early stages of chick embryo development.

    PubMed

    Manca, Annalisa; Capsoni, Simona; Di Luzio, Anna; Vignone, Domenico; Malerba, Francesca; Paoletti, Francesca; Brandi, Rossella; Arisi, Ivan; Cattaneo, Antonino; Levi-Montalcini, Rita

    2012-02-07

    Nerve growth factor (NGF) was discovered because of its neurotrophic actions on sympathetic and sensory neurons in the developing chicken embryo. NGF was subsequently found to influence and regulate the function of many neuronal and non neuronal cells in adult organisms. Little is known, however, about the possible actions of NGF during early embryonic stages. However, mRNAs encoding for NGF and its receptors TrkA and p75(NTR) are expressed at very early stages of avian embryo development, before the nervous system is formed. The question, therefore, arises as to what might be the functions of NGF in early chicken embryo development, before its well-established actions on the developing sympathetic and sensory neurons. To investigate possible roles of NGF in the earliest stages of development, stage HH 11-12 chicken embryos were injected with an anti-NGF antibody (mAb αD11) that binds mature NGF with high affinity. Treatment with anti-NGF, but not with a control antibody, led to a dose-dependent inversion of the direction of axial rotation. This effect of altered rotation after anti NGF injection was associated with an increased cell death in somites. Concurrently, a microarray mRNA expression analysis revealed that NGF neutralization affects the expression of genes linked to the regulation of development or cell proliferation. These results reveal a role for NGF in early chicken embryo development and, in particular, in the regulation of somite survival and axial rotation, a crucial developmental process linked to left-right asymmetry specification.

  1. Single Cycle Structure-Based Humanization of an Anti-Nerve Growth Factor Therapeutic Antibody

    PubMed Central

    Covaceuszach, Sonia; Marinelli, Sara; Krastanova, Ivet; Ugolini, Gabriele; Pavone, Flaminia; Lamba, Doriano; Cattaneo, Antonino

    2012-01-01

    Most forms of chronic pain are inadequately treated by present therapeutic options. Compelling evidence has accumulated, demonstrating that Nerve Growth Factor (NGF) is a key modulator of inflammatory and nociceptive responses, and is a promising target for the treatment of human pathologies linked to chronic and inflammatory pain. There is therefore a growing interest in the development of therapeutic molecules antagonising the NGF pathway and its nociceptor sensitization actions, among which function-blocking anti-NGF antibodies are particularly relevant candidates. In this respect, the rat anti-NGF αD11 monoclonal antibody (mAb) is a potent antagonist, able to effectively antagonize rodent and human NGF in a variety of in vitro and in vivo systems. Here we show that mAb αD11 displays a significant analgesic effect in two different models of persistent pain in mice, with a remarkable long-lasting activity. In order to advance αD11 mAb towards its clinical application in man, anti-NGF αD11 mAb was humanized by applying a novel single cycle strategy based on the a priori experimental determination of the crystal and molecular structure of the parental Fragment antigen-binding (Fab). The humanized antibody (hum-αD11) was tested in vitro and in vivo, showing that the binding mode and the NGF neutralizing biological activities of the parental antibody are fully preserved, with even a significant affinity improvement. The results firmly establish hum-αD11 as a lead candidate for clinical applications in a therapeutic area with a severe unmet medical need. More generally, the single-cycle structure-based humanization method represents a considerable improvement over the standard humanization methods, which are intrinsically empirical and require several refinement cycles. PMID:22403636

  2. Nerve growth factor reduces amiloride‐sensitive Na+ transport in human airway epithelial cells

    PubMed Central

    Shimko, Michael J.; Zaccone, Eric J.; Thompson, Janet A.; Schwegler‐Berry, Diane; Kashon, Michael L.; Fedan, Jeffrey S.

    2014-01-01

    Abstract Nerve growth factor (NGF) is overexpressed in patients with inflammatory lung diseases, including virus infections. Airway surface liquid (ASL), which is regulated by epithelial cell ion transport, is essential for normal lung function. No information is available regarding the effect of NGF on ion transport of airway epithelium. To investigate whether NGF can affect ion transport, human primary air‐interface cultured epithelial cells were placed in Ussing chambers to obtain transepithelial voltage (−7.1 ± 3.4 mV), short‐circuit current (Isc, 5.9 ± 1.0 μA), and transepithelial resistance (750 Ω·cm2), and to measure responses to ion transport inhibitors. Amiloride (apical, 3.5 × 10−5 mol/L) decreased Isc by 55.3%. Apically applied NGF (1 ng/mL) reduced Isc by 5.3% in 5 min; basolaterally applied NGF had no effect. The response to amiloride was reduced (41.6%) in the presence of NGF. K‐252a (10 nmol/L, apical) did not itself affect Na+ transport, but it attenuated the NGF‐induced reduction in Na+ transport, indicating the participation of the trkA receptor in the NGF‐induced reduction in Na+ transport. PD‐98059 (30 μmol/L, apical and basolateral) did not itself affect Na+ transport, but attenuated the NGF‐induced reduction in Na+ transport, indicating that trkA activated the Erk 1/2 signaling cascade. NGF stimulated phosphorylation of Erk 1/2 and the β‐subunit of ENaC. K‐252a and PD‐98059 inhibited these responses. NGF had no effect on Isc in the presence of apical nystatin (50 μmol/L). These results indicate that NGF inhibits Na+ transport through a trkA‐Erk 1/2‐activated signaling pathway linked to ENaC phosphorylation. PMID:25347857

  3. Nerve Growth Factor gene ovarian expression, polymorphism identification, and association with litter size in goats.

    PubMed

    Naicy, T; Venkatachalapathy, R T; Aravindakshan, T V; Radhika, G; Raghavan, K C; Mini, M; Shyama, K

    2016-12-01

    The Nerve Growth Factor (NGF) plays an important role in reproduction by augmenting folliculogenesis. In this study, the coding regions of caprine NGF gene were analyzed to detect single-nucleotide polymorphisms (SNPs), their association with litter size, and the relative ovarian expression of NGF gene in the two indigenous goat breeds of South India viz., the prolific Malabari and less-prolific Attappady Black. The sequence analysis of the third exon containing the entire open reading frame of NGF gene was observed to be of 808 bp with one nonsynonymous mutation at 217th position. Later, polymerase chain reaction (PCR) was performed to amplify a region of 188 bp covering the region carrying the detected mutation. The genomic DNAs from the goats under study (n = 277) were subjected to PCR and single strand conformation polymorphism (SSCP). On analysis, four diplotypes viz., AA, AB, AC, and AD were observed with respective frequencies of 0.50, 0.22, 0.27, and 0.01. Sequencing of the representative samples revealed an additional synonymous mutation, i.e., g.291C>A. Statistical analysis indicated that NGF diplotypes and the SNP g.217G>A were associated with litter size in goats (P < 0.05). Relative expression of NGF gene was significantly higher in the ovaries of goats with history of multiple than single births (P < 0.05). The results of the present study suggest the significant effect of the NGF gene on litter size in goats and identified SNPs would benefit the selection of prolific animals in future marker-assisted breeding programs. The two novel PCR-restriction fragment length polymorphisms designed, based on the detected SNPs, would help in the rapid screening of large number of animals in a breeding population for identifying individual animals with desired genetic characteristics. Copyright © 2016 Elsevier Inc. All rights reserved.

  4. Characterization and localization of nerve growth factor receptors in the embryonic otic vesicle and cochleovestibular ganglion

    SciTech Connect

    Bernd, P.; Represa, J. )

    1989-07-01

    We have investigated the possibility that nerve growth factor (NGF) may play a role in the development of the inner ear. Primordia of the inner ear, the otic vesicle (OV) and cochleovestibular ganglion (CVG), were isolated from 72-hr (stage 19-20) quail embryos and examined for the presence of NGF receptors. Quantitative binding studies revealed that both OV and CVG exhibited specific 125I-NGF binding; levels of nonspecific binding were 6 to 26% of total binding. Scatchard analysis yielded a linear plot, indicating the presence of a single class of NGF receptor. The average binding constant (Kd) was 8.0 nM for OV and 8.6 nM for CVG, corresponding to the low affinity (site II) NGF receptor. Examination of light microscopic radioautographs indicated that most of the specific 125I-NGF binding was located in the ventromedial wall of the OV, with little or no binding in the lateral wall and endolymphatic primordia. These studies were corroborated by microdissection of OV, in which 70% of the radioactivity was found to be localized in the medial half of the OV. In CVG, specific 125I-NGF binding was more concentrated in the cochlear portion of the ganglion, with silver grains primarily over areas containing support cells and immature neurons. Quantitative binding studies with isolated cochlear and vestibular ganglia obtained from 144-hr (stage 29-30) quail embryos revealed that the cochlear ganglion exhibited three times more specific 125I-NGF binding than the vestibular ganglion. The presence of NGF receptors on OV and CVG suggests that these structures are responsive to and/or dependent upon NGF. The following paper examines the question of whether NGF serves either as a mitogen, a survival factor, or a differentiation factor in this system.

  5. Neurosteroid Dehydroepiandrosterone Interacts with Nerve Growth Factor (NGF) Receptors, Preventing Neuronal Apoptosis

    PubMed Central

    Alexaki, Vassilia-Ismini; Avlonitis, Nicolaos; Pediaditakis, Iosif; Efstathopoulos, Paschalis; Calogeropoulou, Theodora; Castanas, Elias; Gravanis, Achille

    2011-01-01

    The neurosteroid dehydroepiandrosterone (DHEA), produced by neurons and glia, affects multiple processes in the brain, including neuronal survival and neurogenesis during development and in aging. We provide evidence that DHEA interacts with pro-survival TrkA and pro-death p75NTR membrane receptors of neurotrophin nerve growth factor (NGF), acting as a neurotrophic factor: (1) the anti-apoptotic effects of DHEA were reversed by siRNA against TrkA or by a specific TrkA inhibitor; (2) [3H]-DHEA binding assays showed that it bound to membranes isolated from HEK293 cells transfected with the cDNAs of TrkA and p75NTR receptors (KD: 7.4±1.75 nM and 5.6±0.55 nM, respectively); (3) immobilized DHEA pulled down recombinant and naturally expressed TrkA and p75NTR receptors; (4) DHEA induced TrkA phosphorylation and NGF receptor-mediated signaling; Shc, Akt, and ERK1/2 kinases down-stream to TrkA receptors and TRAF6, RIP2, and RhoGDI interactors of p75NTR receptors; and (5) DHEA rescued from apoptosis TrkA receptor positive sensory neurons of dorsal root ganglia in NGF null embryos and compensated NGF in rescuing from apoptosis NGF receptor positive sympathetic neurons of embryonic superior cervical ganglia. Phylogenetic findings on the evolution of neurotrophins, their receptors, and CYP17, the enzyme responsible for DHEA biosynthesis, combined with our data support the hypothesis that DHEA served as a phylogenetically ancient neurotrophic factor. PMID:21541365

  6. The potential role of nerve growth factor, brain-derived neurotrophic factor and neurotrophin-3 in avian cochlear and vestibular ganglia development.

    PubMed

    Bernd, P; Zhang, D; Yao, L; Rozenberg, I

    1994-12-01

    The role of the nerve growth factor family of neurotrophins in the development of cochlear and vestibular ganglia is unclear. In order to predict the potential importance of nerve growth factor, brain-derived neurotrophic factor or neurotrophin-3, we examined the expression of neurotrophin mRNA and full-length neurotrophin receptor mRNA by in-situ hybridization and reverse transcription-polymerase chain reaction, as well as whether high affinity 125I-nerve growth factor binding was present, in cochlear and vestibular ganglia of the quail at several stages of development (stages 26, 31 and 36). Nerve growth factor, brain-derived neurotrophic factor and neurotrophin-3 mRNA was detected at all ages examined, suggesting that these neurotrophins may serve an autocrine or paracrine function, especially prior to target contact. In addition, we found full-length trkA and trkC mRNA was expressed, the products of which are the functional neuronal receptors for nerve growth factor and neurotrophin-3, respectively. Although full-length trkA mRNA was found, physiologically important high affinity 125I-nerve growth factor binding was not detected. Since nerve growth factor's effects on survival and neurite outgrowth are mediated through high affinity binding, nerve growth factor may serve an as yet unidentified role in this system. Full-length trkB mRNA, the product of which is the functional neuronal receptor for brain-derived neurotrophic factor, was not detected using reverse transcription-polymerase chain reaction, however, truncated (non-catalytic) trkB was present, at least in cochlear ganglia at stage 31. It is not known what function may be subserved by these truncated receptors.

  7. Control of Growth Within Drosophila Peripheral Nerves by Ras and Protein Kinase A

    DTIC Science & Technology

    2008-02-01

    via induced expression of a nuclear -localized GFP. We also visualized the total complement of peripheral nerve nuclei (peripheral and perineurial glial...RafF179 (Brand and Perrimon, 1994), UAS–green fluorescent pro- tein (GFP) nuclear localization signal (nls) (Shiga et al., 1996), and Akt4226 (Perrimon et... nuclear -localized GFP. We also visualized the total complement of peripheral nerve nuclei (peripheral and perineurial glial) via the Hoechst DNA dye. As

  8. Chronic Nerve Growth Factor Exposure Increases Apoptosis in a Model of In Vitro Induced Conjunctival Myofibroblasts

    PubMed Central

    Micera, Alessandra; Puxeddu, Ilaria; Balzamino, Bijorn Omar; Bonini, Stefano; Levi-Schaffer, Francesca

    2012-01-01

    In the conjunctiva, repeated or prolonged exposure to injury leads to tissue remodeling and fibrosis associated with dryness, lost of corneal transparency and defect of ocular function. At the site of injury, fibroblasts (FB) migrate and differentiate into myofibroblasts (myoFB), contributing to the healing process together with other cell types, cytokines and growth factors. While the physiological deletion of MyoFB is necessary to successfully end the healing process, myoFB prolonged survival characterizes the pathological process of fibrosis. The reason for myoFB persistence is poorly understood. Nerve Growth Factor (NGF), often increased in inflamed stromal conjunctiva, may represent an important molecule both in many inflammatory processes characterized by tissue remodeling and in promoting wound-healing and well-balanced repair in humans. NGF effects are mediated by the specific expression of the NGF neurotrophic tyrosine kinase receptor type 1 (trkANGFR) and/or the pan-neurotrophin glycoprotein receptor (p75NTR). Therefore, a conjunctival myoFB model (TGFβ1-induced myoFB) was developed and characterized for cell viability/proliferation as well as αSMA, p75NTR and trkANGFR expression. MyoFB were exposed to acute and chronic NGF treatment and examined for their p75NTR/trkANGFR, αSMA/TGFβ1 expression, and apoptosis. Both NGF treatments significantly increased the expression of p75NTR, associated with a deregulation of both αSMA/TGFβ1 genes. Acute and chronic NGF exposures induced apoptosis in p75NTR expressing myoFB, an effect counteracted by the specific trkANGFR and/or p75NTR inhibitors. Focused single p75NTR and double trkANGFR/p75NTR knocking-down experiments highlighted the role of p75NTR in NGF-induced apoptosis. Our current data indicate that NGF is able to trigger in vitro myoFB apoptosis, mainly via p75NTR. The trkANGFR/p75NTR ratio in favor of p75NTR characterizes this process. Due to the lack of effective pharmacological agents for balanced

  9. Verticillin A Inhibits Leiomyosarcoma and Malignant Peripheral Nerve Sheath Tumor Growth via Induction of Apoptosis

    PubMed Central

    Zewdu, A; Lopez, G; Braggio, D; Kenny, C; Constantino, D; Bid, HK; Batte, K; Iwenofu, OH; Oberlies, NH; Pearce, CJ; Strohecker, AM; Lev, D; Pollock, RE

    2017-01-01

    Objective The heterogeneity of soft tissue sarcoma (STS) represents a major challenge for the development of effective therapeutics. Comprised of over 50 different histology subtypes of various etiologies, STS subsets are further characterized as either karyotypically simple or complex. Due to the number of genetic anomalies associated with genetically complex STS, development of therapies demonstrating potency against this STS cluster is especially challenging and yet greatly needed. Verticillin A is a small molecule natural product with demonstrated anticancer activity; however, the efficacy of this agent has never been evaluated in STS. Therefore, the goal of this study was to explore verticillin A as a potential STS therapeutic. Methods We performed survival (MTS) and clonogenic analyses to measure the impact of this agent on the viability and colony formation capability of karyotypically complex STS cell lines: malignant peripheral nerve sheath tumor (MPNST) and leiomyosarcoma (LMS). The in vitro effects of verticillin A on apoptosis were investigated through annexin V/PI flow cytometry analysis and by measuring fluorescently-labeled cleaved caspase 3/7 activity. The impact on cell cycle progression was assessed via cytometric measurement of propidium iodide intercalation. In vivo studies were performed using MPNST xenograft models. Tumors were processed and analyzed using immunohistochemistry (IHC) for verticillin A effects on growth (Ki67) and apoptosis (cleaved caspase 3). Results Treatment with verticillin A resulted in decreased STS growth and an increase in apoptotic levels after 24 h. 100 nM verticillin A induced significant cellular growth abrogation after 24 h (96.7, 88.7, 72.7, 57, and 39.7% reduction in LMS1, S462, ST88, SKLMS1, and MPNST724, respectively). We observed no arrest in cell cycle, elevated annexin, and a nearly two-fold increase in cleaved caspase 3/7 activity in all MPNST and LMS cell lines. Control normal human Schwann (HSC) and

  10. Vascular endothelial growth factor gene therapy improves nerve regeneration in a model of obstetric brachial plexus palsy.

    PubMed

    Hillenbrand, Matthias; Holzbach, Thomas; Matiasek, Kaspar; Schlegel, Jürgen; Giunta, Riccardo E

    2015-03-01

    The treatment of obstetric brachial plexus palsy has been limited to conservative therapies and surgical reconstruction of peripheral nerves. In addition to the damage of the brachial plexus itself, it also leads to a loss of the corresponding motoneurons in the spinal cord, which raises the need for supportive strategies that take the participation of the central nervous system into account. Based on the protective and regenerative effects of VEGF on neural tissue, our aim was to analyse the effect on nerve regeneration by adenoviral gene transfer of vascular endothelial growth factor (VEGF) in postpartum nerve injury of the brachial plexus in rats. In the present study, we induced a selective crush injury to the left spinal roots C5 and C6 in 18 rats within 24 hours after birth and examined the effect of VEGF-gene therapy on nerve regeneration. For gene transduction an adenoviral vector encoding for VEGF165 (AdCMV.VEGF165) was used. In a period of 11 weeks, starting 3 weeks post-operatively, functional regeneration was assessed weekly by behavioural analysis and force measurement of the upper limb. Morphometric evaluation was carried out 8 months post-operatively and consisted of a histological examination of the deltoid muscle and the brachial plexus according to defined criteria of degeneration. In addition, atrophy of the deltoid muscle was evaluated by weight determination comparing the left with the right side. VEGF expression in the brachial plexus was quantified by an enzyme-linked immunosorbent assay (ELISA). Furthermore the motoneurons of the spinal cord segment C5 were counted comparing the left with the right side. On the functional level, VEGF-treated animals showed faster nerve regeneration. It was found less degeneration and smaller mass reduction of the deltoid muscle in VEGF-treated animals. We observed significantly less degeneration of the brachial plexus and a greater number of surviving motoneurons (P < 0·05) in the VEGF group. The results of

  11. Expression of neuropeptides and growth-associated protein 43 (GAP-43) in cutaneous and mucosal nerve structures of the adult rat lower lip after mental nerve section.

    PubMed

    Verzè, L; Paraninfo, A; Viglietti-Panzica, C; Panzica, G C; Ramieri, G

    2003-01-01

    The reinnervation of the adult rat lower lip has been investigated after unilateral section of the mental nerve. Rats were sacrificed at 4, 7, 9, 14, 30, and 90 days after the operation. A further group of animals with section of the mental nerve and block of the alveolar nerve regeneration, was sacrificed at 14 days. Specimens were processed for immunocytochemistry with antibodies against PGP 9.5, GAP-43 or neuropeptides (CGRP, SP and VIP). Four days after nerve section, axonal degeneration seems evident in the mental nerve branches and inside skin and mucosa. GAP-43 immunoreactivity is intense in the mental nerve 7 days after nerve section and it reaches its maximal expression and distribution in peripheral nerve fibres at 14 days. At 30 days, the decline in its expression is associated with the increase of PGP9.5-, SP-, and CGRP immunopositivity. VIP is observed only in perivascular fibres at all times observed. Present results suggest that, after sensory denervation of the rat lip, nerve fibres in skin and mucosa remain at lower density than normal. The different time courses in the expression of neuropeptides and GAP-43 suggest a possible early involvement of GAP-43 in peripheral nerve regeneration.