Expression of fibroblast growth factor 10 and its receptor, fibroblast growth factor receptor 2B, in the bovine corpus luteum

There is evidence that several fibroblast growth factors (FGFs) are involved in growth and development of the corpus luteum (CL), but many FGFs have not been investigated in this tissue, including FGF10. The objective of this study was to determine if FGF10 and its receptor (FGFR2B) are expressed in the CL. Bovine CL were collected from an abattoir and classed as corpus hemorrhagica (stage 1), developing (stage 11), developed (stage 111), and regressed (stage IV) CL. Expression of FGF10 and FGFR2B mRNA was measured by reverse transcription-polymerase chain reaction (RT-PCR). Both genes were expressed in bovine CL, and FGF10 expression did not differ between stages of CL development. FGF10 protein was localized to large and small luteal cells by immunohistochemistry. FGFR2B expression was approximately threefold higher in regressed compared to developing and developed CL (P < 0.05). To determine if FGF10 and FGFR2B expression is regulated during functional luteolysis, cattle were injected with PGF2 alpha and CL collected at 0, 0.5, 2, 4, 12, 24, 48, and 64 hr thereafter (n = 5 CL/time point), and mRNA abundance was measured by real-time RT-PCR. FGF10 mRNA expression did not change during functional luteolysis, whereas FGFR2B mRNA abundance decreased significantly at 2, 4, and 12 hr after PGF2a, and returned to pretreatment levels for the period 24-64 hr post-PGF2 alpha. These data suggest a potential role for FGFR2B signaling during structural luteolysis in bovine CL.

Expression and function of fibroblast growth factor 10 and its receptor, fibroblast growth factor receptor 213, in bovine follicles

Some fibroblast growth factors (FGFs) affect ovarian follicle cell growth and/or differentiation. Whereas many FGFs activate several FGF receptors, FGF7 and FGF10 primarily activate only one, FGFR2B. As FGF7 is produced by bovine theca cells and acts on granulosa cells, we tested the hypothesis that FGF10 may also play a role in folliculogenesis in cattle. Reverse transcription-polymerase chain reaction demonstrated the presence of FGF10 mRNA in the oocytes and theca cells of the antral follicles, as well as in the preantral follicles. FGF10 protein was detected by immunohistochemistry in the oocytes of the preantral and antral follicles, and in the granulosa and theca cells of the antral follicles. FGF10 expression in theca cells changed during follicle development; mRNA abundance decreased with increasing follicular estradiol concentration in healthy follicles, and was lowest in highly atretic follicles. Culturing of granulosa cells in serum-free medium revealed FSH regulation of FGF10 receptor expression. The addition of FGF10 to cultured granulosa cells decreased the level of estradiol but did not alter cell proliferation. These data support a role for FGF10 in signaling to granulosa cells from theca cells and/or the oocyte.

Physical exercise modulates peripheral levels of brain-derived neurotrophic factor (BDNF): A systematic review of experimental studies in the elderly

The objective of this study was to conduct a systematic review of studies that analyzed the effect of physical exercise on the peripheral levels of BDNF in elderly individuals. Method: We conducted a search in PsycINFO, Biological Abstracts, Pubmed, Web of Science, and Science Direct from 1990 to 2011, using the following keywords: physical exercise , physical activity , physical therapy , training , BDNF , neuroplasticity , neurotrophins , neuroplasticity proteins , aged , older , elderly The articles were considered for inclusion in the review if they were studies with elderly, assessed peripheral (serum and/or plasma) BDNF and evaluated an acute exercise or chronic exercise (training). Results: Five randomized controlled trial and one randomized non-controlled trial studies were analyzed. Five out of six studies reported a significantly higher BDNF response to aerobic acute exercise and to aerobic or strength training program in healthy elderly and elderly with different pathologies. Conclusion: It was not possible to establish a recommendation protocol for the type and intensity of physical exercise required to produce an increase in levels BDNF. However, physical exercise, particularly, moderate-intensity exercises seem to be more effective to promote increase the peripheral levels of BDNF in the elderly. © 2012 Elsevier B.V.

Acute Aerobic Exercise Increases Brain-Derived Neurotrophic Factor Levels in Elderly with Alzheimer's Disease

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Regulación de la proliferación tumoral y la apoptosis en tumores epidermoides de cavidad oral: papel del receptor para el factor de crecimiento insulínico tipo 1 (IGF-1R) y major vault proteína (MVP)

Programa de doctorado: Cáncer: Biología y Clínica

Effects of local continuous release of brain derived neurotrophic factor (BDNF) on peripheral nerve regeneration in a rat model

The purpose of this study was to evaluate the effect of continuously released BDNF on peripheral nerve regeneration in a rat model. Initial in vitro evaluation of calcium alginate prolonged-release-capsules (PRC) proved a consistent release of BDNF for a minimum of 8 weeks. In vivo, a worst case scenario was created by surgical removal of a 20-mm section of the sciatic nerve of the rat. Twenty-four autologous fascia tubes were filled with calcium alginate spheres and sutured to the epineurium of both nerve ends. The animals were divided into 3 groups. In group 1, the fascial tube contained plain calcium alginate spheres. In groups 2 and 3, the fascial tube contained calcium alginate spheres with BDNF alone or BDNF stabilized with bovine serum albumin, respectively. The autocannibalization of the operated extremity was clinically assessed and documented in 12 additional rats. The regeneration was evaluated histologically at 4 weeks and 10 weeks in a blinded manner. The length of nerve fibers and the numbers of axons formed in the tube was measured. Over a 10-week period, axons have grown significantly faster in groups 2 and 3 with continuously released BDNF compared to the control. The rats treated with BDNF (groups 2 and 3) demonstrated significantly less autocannibalization than the control group (group 1). These results suggest that BDNF may not only stimulate faster peripheral nerve regeneration provided there is an ideal, biodegradable continuous delivery system but that it significantly reduces the neuropathic pain in the rat model.

Brain-derived neurotrophic factor protects against multiple forms of brain injury in bacterial meningitis

BACKGROUND Brain-derived neurotrophic factor (BDNF) blocks activation of caspase-3, reduces translocation of apoptosis-inducing factor (AIF), attenuates excitotoxicity of glutamate, and increases antioxidant enzyme activities. The mechanisms of neuroprotection suggest that BDNF may be beneficial in bacterial meningitis. METHODS To assess a potentially beneficial effect of adjuvant treatment with BDNF in bacterial meningitis, 11-day-old infant rats with experimental meningitis due to Streptococcus pneumoniae or group B streptococci (GBS) were randomly assigned to receive intracisternal injections with either BDNF (3 mg/kg) or equal volumes (10 mu L) of saline. Twenty-two hours after infection, brains were analyzed, by histomorphometrical examination, for the extent of cortical and hippocampal neuronal injury. RESULTS Compared with treatment with saline, treatment with BDNF significantly reduced the extent of 3 distinct forms of brain cell injury in this disease model: cortical necrosis in meningitis due to GBS (median, 0.0% [range, 0.0%-33.7%] vs. 21.3% [range, 0.0%-55.3%]; P<.03), caspase-3-dependent cell death in meningitis due to S. pneumoniae (median score, 0.33 [range, 0.0-1.0] vs. 1.10 [0.10-1.56]; P<.05), and caspase-3-independent hippocampal cell death in meningitis due to GBS (median score, 0 [range, 0-2] vs. 0.88 [range, 0-3.25]; P<.02). The last form of injury was associated with nuclear translocation of AIF. CONCLUSION BDNF efficiently reduces multiple forms of neuronal injury in bacterial meningitis and may hold promise as adjunctive therapy for this disease.

Human NR5A1/SF-1 mutations show decreased activity on BDNF (brain-derived neurotrophic factor), an important regulator of energy balance: testing impact of novel SF-1 mutations beyond steroidogenesis

CONTEXT Human NR5A1/SF-1 mutations cause 46,XY disorder of sex development (DSD) with broad phenotypic variability, and rarely cause adrenal insufficiency although SF-1 is an important transcription factor for many genes involved in steroidogenesis. In addition, the Sf-1 knockout mouse develops obesity with age. Obesity might be mediated through Sf-1 regulating activity of brain-derived neurotrophic factor (BDNF), an important regulator of energy balance in the ventromedial hypothalamus. OBJECTIVE To characterize novel SF-1 gene variants in 4 families, clinical, genetic and functional studies were performed with respect to steroidogenesis and energy balance. PATIENTS 5 patients with 46,XY DSD were found to harbor NR5A1/SF-1 mutations including 2 novel variations. One patient harboring a novel mutation also suffered from adrenal insufficiency. METHODS SF-1 mutations were studied in cell systems (HEK293, JEG3) for impact on transcription of genes involved in steroidogenesis (CYP11A1, CYP17A1, HSD3B2) and in energy balance (BDNF). BDNF regulation by SF-1 was studied by promoter assays (JEG3). RESULTS Two novel NR5A1/SF-1 mutations (Glu7Stop, His408Profs*159) were confirmed. Glu7Stop is the 4th reported SF-1 mutation causing DSD and adrenal insufficiency. In vitro studies revealed that transcription of the BDNF gene is regulated by SF-1, and that mutant SF-1 decreased BDNF promoter activation (similar to steroid enzyme promoters). However, clinical data from 16 subjects carrying SF-1 mutations showed normal birth weight and BMI. CONCLUSIONS Glu7Stop and His408Profs*159 are novel SF-1 mutations identified in patients with 46,XY DSD and adrenal insufficiency (Glu7Stop). In vitro, SF-1 mutations affect not only steroidogenesis but also transcription of BDNF which is involved in energy balance. However, in contrast to mice, consequences on weight were not found in humans with SF-1 mutations.

Neurotrophic factors [activity-dependent neurotrophic factor (ADNF) and basic fibroblast growth factor (bFGF)] interrupt excitotoxic neurodegenerative cascades promoted by a PS1 mutation

Although an excitotoxic mechanism of neuronal injury has been proposed to play a role in chronic neurodegenerative disorders such as Alzheimer’s disease, and neurotrophic factors have been put forward as potential therapeutic agents, direct evidence is lacking. Taking advantage of the fact that mutations in the presenilin-1 (PS1) gene are causally linked to many cases of early-onset inherited Alzheimer’s disease, we generated PS1 mutant knock-in mice and directly tested the excitotoxic and neurotrophic hypotheses of Alzheimer’s disease. Primary hippocampal neurons from PS1 mutant knock-in mice exhibited increased production of amyloid β-peptide 42/43 and increased vulnerability to excitotoxicity, which occurred in a gene dosage-dependent manner. Neurons expressing mutant PS1 exhibited enhanced calcium responses to glutamate and increased oxyradical production and mitochondrial dysfunction. Pretreatment with either basic fibroblast growth factor or activity-dependent neurotrophic factor protected neurons expressing mutant PS1 against excitotoxicity. Both basic fibroblast growth factor and activity-dependent neurotrophic factor stabilized intracellular calcium levels and abrogated the increased oxyradical production and mitochondrial dysfunction otherwise caused by the PS1 mutation. Our data indicate that neurotrophic factors can interrupt excitotoxic neurodegenerative cascades promoted by PS1 mutations.

Intrastriatal injection of an adenoviral vector expressing glial-cell-line-derived neurotrophic factor prevents dopaminergic neuron degeneration and behavioral impairment in a rat model of Parkinson disease

Glial-cell-line-derived neurotrophic factor (GDNF) is a potent neurotrophic factor for adult nigral dopamine neurons in vivo. GDNF has both protective and restorative effects on the nigro-striatal dopaminergic (DA) system in animal models of Parkinson disease. Appropriate administration of this factor is essential for the success of its clinical application. Since it cannot cross the blood–brain barrier, a gene transfer method may be appropriate for delivery of the trophic factor to DA cells. We have constructed a recombinant adenovirus (Ad) encoding GDNF and injected it into rat striatum to make use of its ability to infect neurons and to be retrogradely transported by DA neurons. Ad-GDNF was found to drive production of large amounts of GDNF, as quantified by ELISA. The GDNF produced after gene transfer was biologically active: it increased the survival and differentiation of DA neurons in vitro. To test the efficacy of the Ad-mediated GDNF gene transfer in vivo, we used a progressive lesion model of Parkinson disease. Rats received injections unilaterally into their striatum first of Ad and then 6 days later of 6-hydroxydopamine. We found that mesencephalic nigral dopamine neurons of animals treated with the Ad-GDNF were protected, whereas those of animals treated with the Ad-β-galactosidase were not. This protection was associated with a difference in motor function: amphetamine-induced turning was much lower in animals that received the Ad-GDNF than in the animals that received Ad-β-galactosidase. This finding may have implications for the development of a treatment for Parkinson disease based on the use of neurotrophic factors.

Glial cell line-derived neurotrophic factor promotes the development of adrenergic neurons in mouse neural crest cultures

Growth of mouse neural crest cultures in the presence of glial cell line-derived neurotrophic factor (GDNF) resulted in a dramatic dose-dependent increase in the number of tyrosine hydroxylase (TH)-positive cells that developed when 5% chicken embryo extract was present in the medium. In contrast, growth in the presence of bone morphogenetic protein (BMP)-2, BMP-4, BMP-6, transforming growth factor (TGF) β1, TGF-β2, and TGF-β3 elicited no increase in the number of TH-positive cells. The TH-positive cells that developed in the presence of GDNF had neuronal morphology and contained the middle and low molecular weight neurofilament proteins. Numerous TH-negative cells with the morphology of neurons also were observed in GDNF-treated cultures. Analysis revealed that the period from 6 to 12 days in vitro was the critical time for exposure to GDNF to generate the increase in TH-positive cell number. The growth factors neurotrophin-3 and fibroblast growth factor-2 elicited increases in the number of TH-positive cells similar to that seen in response to GDNF. In contrast, nerve growth factor was unable to substitute for GDNF. These findings extend the previously reported biological activities of GDNF by showing that it can act on mouse neural crest cultures to promote the development of neurons.

Brain-derived neurotrophic factor-deficient mice develop aggressiveness and hyperphagia in conjunction with brain serotonergic abnormalities

Brain-derived neurotrophic factor (BDNF) has trophic effects on serotonergic (5-HT) neurons in the central nervous system. However, the role of endogenous BDNF in the development and function of these neurons has not been established in vivo because of the early postnatal lethality of BDNF null mice. In the present study, we use heterozygous BDNF+/− mice that have a normal life span and show that these animals develop enhanced intermale aggressiveness and hyperphagia accompanied by significant weight gain in early adulthood; these behavioral abnormalities are known to correlate with 5-HT dysfunction. Forebrain 5-HT levels and fiber density in BDNF+/− mice are normal at an early age but undergo premature age-associated decrements. However, young adult BDNF+/− mice show a blunted c-fos induction by the specific serotonin releaser-uptake inhibitor dexfenfluramine and alterations in the expression of several 5-HT receptors in the cortex, hippocampus, and hypothalamus. The heightened aggressiveness can be ameliorated by the selective serotonin reuptake inhibitor fluoxetine. Our results indicate that endogenous BDNF is critical for the normal development and function of central 5-HT neurons and for the elaboration of behaviors that depend on these nerve cells. Therefore, BDNF+/− mice may provide a useful model to study human psychiatric disorders attributed to dysfunction of serotonergic neurons.

Tissue factor- and factor X-dependent activation of protease-activated receptor 2 by factor VIIa

Protease-activated receptor 2 (PAR2) is expressed by vascular endothelial cells and other cells in which its function and physiological activator(s) are unknown. Unlike PAR1, PAR3, and PAR4, PAR2 is not activatable by thrombin. Coagulation factors VIIa (FVIIa) and Xa (FXa) are proteases that act upstream of thrombin in the coagulation cascade and require cofactors to interact with their substrates. These proteases elicit cellular responses, but their receptor(s) have not been identified. We asked whether FVIIa and FXa might activate PARs if presented by their cofactors. Co-expression of tissue factor (TF), the cellular cofactor for FVIIa, together with PAR1, PAR2, PAR3, or PAR4 conferred TF-dependent FVIIa activation of PAR2 and, to lesser degree, PAR1. Responses to FXa were also observed but were independent of exogenous cofactor. The TF/FVIIa complex converts the inactive zymogen Factor X (FX) to FXa. Strikingly, when FX was present, low picomolar concentrations of FVIIa caused robust signaling in cells expressing TF and PAR2. Responses in keratinocytes and cytokine-treated endothelial cells suggested that PAR2 may be activated directly by TF/FVIIa and indirectly by TF/FVIIa-generated FXa at naturally occurring expression levels of TF and PAR2. These results suggest that PAR2, although not activatable by thrombin, may nonetheless function as a sensor for coagulation proteases and contribute to endothelial activation in the setting of injury and inflammation. More generally, these findings highlight the potential importance of cofactors in regulating PAR function and specificity.