Nerve Growth Factor Regulation Research Articles

Nerve growth factor in the synovial fluid of patients with chronic arthritis.

Cytokines regulate nerve growth factor (NGF) synthesis during inflammatory processes. Since cytokines are also involved in the inflammatory processes of autoimmune rheumatic diseases, we examined levels of NGF in patients with rheumatoid or other types of chronic arthritis. NGF was present in the synovial fluid and synovium of patients with chronic arthritis, but was undetectable in control fluids. We conclude that NGF might be involved in the pathogenesis of arthritis.

Interleukin-1 beta and tumor necrosis factor-alpha synergistically stimulate nerve growth factor synthesis in rat mesangial cells

Recent evidence indicates that cytokines are potent inducers of nerve growth factor (NGF) expression both in peripheral tissues and the central nervous system and that NGF, in addition to its neurotrophic action, also acts as an immunoregulatory agent. It was of interest to investigate whether inflammatory cytokines affect NGF production in renal mesangial cells, which play a crucial role in the modulation of the local immune function in the glomerulus. Our results show that the simultaneous addition of interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) elicited a marked (13-fold) increase of NGF protein released by cultured rat glomerular mesangial cells within 24 h, whereas IL-1 alpha in combination with TNF-alpha, as well as the cytokines alone, did not promote the synthesis of NGF. The synergistic effect was dose dependent (maximal at 1 nM) and due to enhanced gene expression, since the cytokine treatment caused a fivefold increase in NGF mRNA after 8 h. Stimulation of NGF synthesis was abolished by mepacrine and dexamethasone, indicating that phospholipase A2 may be involved in NGF regulation. Moreover, pretreatment of the cells with the lipoxygenase inhibitor nordihydroguaiaretic acid (NDGA) abolished induction of NGF by cytokines; in contrast, the specific cyclooxygenase inhibitors indomethacin and diclofenac failed to modify NGF production. These data suggest that a lipoxygenase metabolite produced in response to IL-1 beta and TNF-alpha acts as a mediator in NGF gene expression. In conclusion, these findings support a model in which a cytokine cascade including NGF may play an important role in the pathophysiology of inflammatory renal diseases.

Depolarizing stimuli regulate nerve growth factor gene expression in cultured hippocampal neurons.

Although trophic factors and neuronal activity have been implicated in regulating functional synaptic circuits, the relationship of trophic interaction to impulse activity in synaptogenesis remains unclear. Using cultured hippocampus as a model system, we provide direct evidence that depolarization and impulse activity specifically increase nerve growth factor gene expression in neurons. Depolarizing stimuli, such as a high K+ concentration or the Na+ channel agonist veratridine, elicited a 3-fold increase of nerve growth factor mRNA levels in both explant and dissociated cultures. Blockade of depolarization by tetrodotoxin prevented the increase of neuronal nerve growth factor mRNA. Further, nerve growth factor gene expression was stimulated by picrotoxin, a gamma-aminobutyric acid antagonist frequently used to enhance hippocampal neuronal activity. Impulse regulation of trophic gene function may be relevant to developmental synaptogenesis and synaptic strengthening in learning and memory.

Kainic acid-induced seizures stimulate increased expression of nerve growth factor mRNA in rat hippocampus

The influence of kainic acid (KA)-induced limbic seizure activity on the expression of mRNA for nerve growth factor (NGF) in adult rat brain was studied using in situ hybridization and S1 nuclease protection techniques with RNA probes complementary to murine and rat NGF mRNA. Within hippocampus, intracerebroventricular injection of 0.5 μg KA caused a dramatic bilateral increase in hybridization of the 35S-labeled cRNA within stratum granulosum. This increase was first evident 1 h post-KA, appeared maximal at approximately 20-fold control levels at 2–3 h post-injection, and declined to control levels by 48 h post-injection. During the period of maximal hybridization, all but the deepest cells within stratum granulosum appeared to be autoradiographically labeled. Hybridization of the NGF cRNA probe was also increased within superficial layers of piriform and entorhinal cortex and, to much lesser extent, within scattered neurons of layers II and III of neocortex in KA-treated rats. In olfactory cortical areas, hybridization was maximally elevated 15.5–24.5 h after KA injection. In contrast to these effects, KA treatment did not consistently influence the density of hybridization, or number of neurons labeled, within the dentate gyrus hilus or the hippocampus proper (CA1–CA3). In agreement with the in situ hybridization results, S1 nuclease protection assay detected KA-induced increases in hybridization within pooled dentate gyrus/CA1 samples, but not hippocampal CA3 samples. These data support the conclusion that seizure activity stimulates a transient increase in NGF expression by select populations of forebrain neurons and indicates that experimental seizure paradigms might be further exploited for analyses of the mechanisms of NGF regulation and processing in the adult brain.

Regulation of schwann cell surface and truncated nerve growth factor receptors in vitro by axonal components

A tissue culture model was designed to characterize the ability of axonal membranes and detergent extracts of membranes to down regulate nerve growth factor (NGF) receptors on the Schwann cell surface. The findings establish that (i) axons from intact neurons, as well as axonal membrane fragments, and extracts of these axons can down regulate Schwann cell NGF receptor number, (ii) central, as well as peripheral, axons have the ability to affect Schwann cell NGF receptor expression, (iii) cross-species reactivity is evident with this phenomenon, (iv) physical contact is necessary for axons to down regulate Schwann cell NGF receptor, (v) the response of axons is glial specific, and (vi) soluble, truncated NGF receptors elaborated by Schwann cells are also down-regulated by axon components.

Glucocorticoid Hormones Negatively Regulate Nerve Growth Factor Expression In Vivo and in Cultured Rat Fibroblasts

Sciatic nerve transection leads to an up-regulation of nerve growth factor (NGF) production in non-neuronal cells surrounding the axons. The lesion-mediated increase in NGF-mRNA levels in the nerve can be blocked by pretreating the animals with the synthetic glucocorticoid dexamethasone. Dexamethasone also reduces NGF-mRNA levels in cultured sciatic fibroblasts stimulated with fetal calf serum or interleukin-1. In order to study at which level glucocorticoids down-regulate NGF expression, sciatic fibroblasts where transfected with a construct in which a reporter gene (chloramphenicol acetyltransferase) is expressed under the control of the NGF promotor. The results demonstrated that dexamethasone effectively represses NGF gene transcription. Deletion experiments showed that a 162 nucleotide promotor region mediates the glucocorticoid hormone suppression of NGF expression. The negative regulation of NGF synthesis by glucocorticoids is a factor to be considered in the treatment of patients with peripheral nerve lesions.

Density-Dependent Nerve Growth Factor Regulation of Gs-α RNA in Pheochromocytoma 12 Cells

Nerve growth factor (NGF) affects levels of the alpha subunit of the stimulatory G protein (Gs-alpha) in pheochromocytoma 12 cells in a bidirectional, density-dependent manner. Cells grown at high density responded to NGF treatment with increased levels of Gs-alpha mRNA and protein. Conversely, in cells grown in low-density cultures, levels of this mRNA were lowered by NGF treatment.

Density-dependent nerve growth factor regulation of Gs-alpha RNA in pheochromocytoma 12 cells.

Nerve growth factor (NGF) affects levels of the alpha subunit of the stimulatory G protein (Gs-alpha) in pheochromocytoma 12 cells in a bidirectional, density-dependent manner. Cells grown at high density responded to NGF treatment with increased levels of Gs-alpha mRNA and protein. Conversely, in cells grown in low-density cultures, levels of this mRNA were lowered by NGF treatment.

Nerve growth factor regulates tyrosine hydroxylase gene transcription through a nucleoprotein complex that contains c-Fos.

We have studied nerve growth factor (NGF) regulation of the expression of the tyrosine hydroxylase (TH) gene in PC12 cells. The TH gene encodes the initial and rate-limiting enzyme of the catecholamine biosynthetic pathway. We show that the TH gene is transiently transcriptionally induced by a mechanism reliant on new protein synthesis during 1-2 hr of NGF stimulation, a time following the induction of the c-fos gene at 15 min post-NGF treatment. A potential regulatory sequence located within the TH gene promoter, the TH-FSE, shares homology to a known regulatory element, the fat-specific element (FSE), which is found upstream from genes activated during adipocyte differentiation and binds the Fos-Jun transcription factor complex. We show that the TH-FSE DNA sequence elevates the basal level of transcription from the rat TH promoter and is required for NGF inducibility. This DNA element binds authentic Fos-Jun products produced abundance during NGF stimulation and by in vitro translation. We demonstrate further that the TH-FSE can bind proteins present in PC12 nuclear extracts in a sequence-specific manner. The DNA/nucleoprotein complex that forms increases in abundance during NGF stimulation and reaches a maximum level at 4 hr of treatment. Antibody inhibition studies utilizing an anti-Fos antibody indicate that Fos and/or Fos-related antigen(s) associate with the TH-FSE and suggest that the Fos protein family contributes to the regulation of TH in vivo. These results support a model in which NGF-induced immediate early genes, including c-Fos, contribute to the regulation of delayed early genes such as TH and thereby control neuronal differentiation.

Nerve growth factor regulates expression of neuropeptide genes in adult sensory neurons.

Nerve growth factor (NGF) is a trophic molecule essential for the survival of sympathetic and sensory neurons during ontogeny. The extent to which NGF is involved in the maintenance or regulation of the differentiated phenotypes of mature peripheral neurons is much less clear, however. Biochemical analysis of the actions of NGF upon peripheral neurons has been hampered by the lack of a preparation of neuronal cells that are responsive to NGF but do not require it for survival. We report here that in adult dorsal root ganglion neurons, which can be isolated, enriched and maintained in culture in the absence of neuronal growth factors, the expression of mRNAs encoding the precursors of two neuropeptides, substance P and calcitonin gene-related peptide is regulated by NGF. Our results provide the first direct evidence of a continuous dynamic role for NGF in regulation of peptide neurotransmitter/neuromodulator levels in mature sensory neurons.

Early polyamine treatment enhances survival of sympathetic neurons after postnatal axonal injury or immunosympathectomy

We have recently demonstrated that following injury of their axon, sympathetic neurons of the rat superior cervical ganglion become dependent on polyamine synthesis for their survival. In addition we have observed that the treatment of newborn rats with biogenic polyamines can prevent the naturally occurring reduction in the number of neurons in the ganglion. In the present study groups of newborn rats were subjected to either postganglionic nerve crush (axotomy) or to treatment with antiserum to nerve growth factor (immunosympathectomy), two treatments which result in a massive loss of neurons in the ganglion. Daily injections of the polyamines putrescine, spermidine and spermine (10 mg/kg each), for 7 days after the operation to the axotomized group, and for 9 days starting with the first antiserum injection to the immunosympathectomized group, attenuated the nerve cell loss. The polyamine treatment also attenuated the reduction in the activity of the neurotransmitter-synthesizing enzyme tyrosine hydroxylase observed after both axotomy and immunosympathectomy in the ganglion. These results further indicate that polyamines are important for the survival of sympathetic neurons and, while their mechanism of action is unknown, an interaction with nerve growth factor regulation cannot be excluded. In the iris, the reduction observed in [ 3H]norepinephrine uptake after the two noxious treatments was unproportionately small when compared to the large drop in the number of parent neurons in the ganglion. This suggests that compensatory mechanisms exist which act to adjust the number of functional axon terminals per neuron so that the number of terminals innervating the target remains relatively constant.

Regulation of Cell Growth

At this meeting of the RSM's Section of Pathology, the regulation of haemopoietic stem cells and growth factors regulating various cell lines were described, and the role of oncogenes, platelet-derived growth factor and nerve growth factor in growth regulation was discussed.

Serum and thyroid hormones T3 and T4 regulate nerve growth factor mRNA levels in mouse L cells

Mouse L cells synthesize and secrete a neurotrophic factor related to the β subunit of the submaxillary gland nerve growth factor (NGF) of male mice. Use of a cDNA probe which encodes the β-NGF mRNA demonstrated that L cells produce a transcript identical in size to that of the submaxillary gland. Moreover, target sites of restriction enzymes EcoRI, PstI and BamHI were not significantly rearranged in the β-NGF gene locus of these cells. The abundance of the β-NGF transcript was found to depend on culture conditions. Removal of serum depressed the cellular content of polyadenylated RNA by a factor of 1.7, and decreased specifically the pool of β-NGF transcript by an additional factor of 4. The presence of 10 −7 M testosterone in the serum-free medium did not modify the level of β-NGF mRNA, while addition of 10 −7 M T3 (or T4) increased this level by a factor of 1.5. These data provide the first evidence that the β-NGF mRNA of L cells is subjected to regulation, but in a way apparently different from that described for the submaxillary gland.

Involvement of Nerve Growth Factor in Neuroendocrine Regulation

Nerve Growth Factor (NGF), a poly-peptide which is essential for development and maintenance of peripheral sympathetic and sensory neurons, has been investigated for its effects on the hypothalamopituitary adrenocortical axis (HPA). Intravenous injection of NGF in rats is followed by a prolonged elevation of plasma-ACTH, betalipotropin, beta-endorphin and corticosterone. In contrast to the acute intensive drinking response, this endocrine effect is independent from the renin contaminant of the standard 2.5 S NGF-preparation. This is supported by the fact, that intraventricular captopril (a potent inhibitor of the angiotensin converting enzyme) abolished the drinking response, but did not block the stimulation of the HPA-axis. In addition a renin-free NGF, prepared by isoelectric focussing, stimulated ACTH-release, but failed to induce thirst.

Sodium dependence of the nerve growth factor — regulated hexose uptake in chick embryo ganglionic cells

Embryonic dorsal root ganglionic cells, when incubated in vitro in the absence of nerve growth factor (NGF) undergo a general metabolic degeneration which is preceded by certain changes in permeation properties. Previous studies demonstrated that NGF can rapidly modulate permeation properties which regulate the availability to the cell of an important energy source, glucose. Hexose uptake was determined by measuring the ability of the cells to accumulate [ 3H]labelled 2-deoxy- d-glucose. The work reported here shows that the NGF-dependent portion (about one-third) of the total specific hexose uptake was also dependent on the presence of Na +, with the apparent uptake constant ( K t ) for deoxyglucose varying inversely with an external Na + concentration of 70–140 mM; V max was unaffected in this range. Preincubation of ganglionic cells with 10 mM ouabain for 15–60 min, followed by a pulse with [ 3H]-deoxyglucose, also resulted in 50–95% reduction of the NGF-sensitive uptake. A similar pretreatment of cells with veratridine gave a 25–50% reduction in uptake. The NGF-controlled hexose uptake was also energy dependent, being diminished 50–95% after a 30–90 min preincubation with 2 mM 2,4-dinitrophenol. Uptake activities for other substrates (α-aminoisobutyric acid, uridine) which exhibited NGF regulation were likewise Na +-sensitive. These results indicate that availability of major energy substrates to NGF-dependent dorsal root ganglionic neurons is controlled by sodium gradients across their membranes. It is conceivable that NGF provides for maintenance and development of its target neurons by acting on such sodium gradients and, consequently, regulating the intake of essential nutrients.