RESUMO
Attention Deficit Hyperactivity Disorder (ADHD) is a neurodevelopmental disorder that presents sex differences in the severity and presentation of symptoms, whose neurobiological basis is still unknown. Both Growth-associated Protein 43 (GAP-43) and Sonic hedgehog (Shh) are considered essential proteins for the appropriate brain development, but their participation in ADHD neurobiology have not been investigated yet. In this study, we hypothesized that alterations in these proteins could be related to behavioral traits to ADHD phenotype. Thus, both sexes of infant Spontaneously hypertensive rats (SHR, used as ADHD animal model) were evaluated for developmental milestones, locomotor activity, olfactory and recognition memory. Both GAP-43 and Shh were assessed in the olfactory bulb, frontal cortex and hippocampus in early and late infancy. During early infancy, SHR reached three developmental milestones later, and females showed olfactory memory impairment accompanied by increased levels of Shh in the olfactory bulb. In later infancy, hyperlocomotion, impaired recognition memory, and decreased Shh in the hippocampus were observed in SHR from both sexes. While in early infancy GAP-43 was not altered, it was decreased in the frontal cortex and hippocampus of female SHR in late infancy. Therefore, both Shh and GAP-43 are involved in the sex-dependent behavioral alterations showed by infant SHR. Despite the disorder's complexity and heterogeneity, our findings reveal important developmental parameters during SHR development and also emphasizes the relevance of studying sex differences in the ADHD context.
Assuntos
Transtorno do Deficit de Atenção com Hiperatividade , Proteínas Hedgehog , Animais , Encéfalo/metabolismo , Modelos Animais de Doenças , Feminino , Proteína GAP-43/metabolismo , Proteínas Hedgehog/metabolismo , Masculino , Transtornos da Memória/metabolismo , Odorantes , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Caracteres SexuaisRESUMO
OBJECTIVE: Malformations of the polymicrogyria spectrum can be mimicked in rodents through neonatal transcranial focal cortical freeze lesions. The animals presenting the malformations present both altered synaptic events and epileptiform activity in the vicinity of the microgyrus, but the comprehension of their contribution to increased predisposition or severity of seizures require further studies. METHODS: In order to investigate these issues, we induced both microgyria and schizencephaly in 57 mice and evaluated: their convulsive susceptibility and severity after pentyleneterazol (PTZ) treatment, the quantification of their symmetric and asymmetric synapses, the morphology of their dendritic arbors, and the content of modulators of synaptogenesis, such as SPARC, gephyrin and GAP-43 within the adjacent visual cortex. RESULTS: Our results have shown that only schizencephalic animals present increased convulsive severity. Nevertheless, both microgyric and schizencephalic cortices present increased synapse number and dendritic complexity of layer IV and layer V-located neurons. Specifically, the microgyric cortex presented reduced inhibitory synapses, while the schizencephalic cortex presented increased excitatory synapses. This altered synapse number is correlated with decreased content of both the anti-synaptogenic factor SPARC and the inhibitory postsynaptic organizer gephyrin in both malformed groups. Besides, GAP-43 content and dendritic spines number are enhanced exclusively in schizencephalic cortices. SIGNIFICANCE: In conclusion, our study supports the hypothesis that the sum of synaptic alterations drives to convulsive aggravation in animals with schizencephaly, but not microgyria after PTZ treatment. These findings reveal that different malformations of cortical development should trigger epilepsy via different mechanisms, requiring further studies for development of specific therapeutic interventions.
Assuntos
Neocórtex , Polimicrogiria , Esquizencefalia , Animais , Modelos Animais de Doenças , Proteína GAP-43 , Camundongos , Pentilenotetrazol , Convulsões/induzido quimicamenteRESUMO
Neurodegenerative diseases are characterized by progressive loss of the structure and function of specific neuronal populations, and have been associated with reduced neurotrophic support. Neurotrophins, like NGF (nerve growth factor), are endogenous proteins that induce neuritogenesis and modulate axonal growth, branching, and synapsis; however, their therapeutic application is limited mainly by low stability, short half-life, and inability to cross the blood-brain barrier (BBB). Small neurotrophic molecules that have suitable pharmacokinetics and are able to cross the BBB are potential candidates for neuroprotection. Baccharin is a bioactive small molecule isolated from Brazilian green propolis. In the present study, we investigated the neurotrophic and neuroprotective potential of baccharin in the PC12 cell neuronal model. We used pharmacological inhibitors (K252a, LY294002, and U0126), and ELISA (phospho-trkA, phospho-Akt, and phospho-MEK) to investigate the involvement of trkA receptor, PI3k/Akt pathway, and MAPK/Erk pathway, respectively. Additionally, we evaluated the expression of axonal (GAP-43) and synaptic (synapsin I) proteins by western blot. The results showed that baccharin induces neuritogenesis in NGF-deprived PC12 cells, through activation of trkA receptor and the downstream signaling cascades (PI3K/Akt and MAPK/ERK), which is the same neurotrophic pathway activated by NGF in PC12 cells and neurons. Baccharin also induced the expression of GAP-43 and synapsin I, which mediate axonal and synaptic plasticity, respectively. Additionally, in silico predictions of baccharin showed favorable physicochemical properties, pharmacokinetics, drug-likeness, and medicinal chemistry friendliness. Altogether, these findings suggest that baccharin is a promising neurotrophic agent whose therapeutic application in neurodegeneration should be further investigated.
Assuntos
Fator de Crescimento Neural , Própole , Animais , Brasil , Proteína GAP-43/metabolismo , Fator de Crescimento Neural/metabolismo , Fator de Crescimento Neural/farmacologia , Células PC12 , Fosfatidilinositol 3-Quinases/metabolismo , Própole/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Receptor trkA/metabolismo , Transdução de Sinais , Sinapsinas/metabolismo , TricotecenosRESUMO
Doxycycline has been used as antibiotic since the 1960s. Recently, studies have shown that doxycycline is neuroprotective in models of neurodegenerative diseases and brain injuries, mainly due to anti-inflammatory and anti-apoptotic effects. However, it is not known if doxycycline has neurotrophic potential, which is relevant, considering the role of axonal degeneration at the early stages of neurodegeneration in Alzheimer's disease, Amyotrophic Lateral Sclerosis and Parkinson's disease as well as in normal aging. Axons are preceded by the formation of neurites, the hallmark of the neuronal differentiation induced by neurotrophins like NGF. Therefore, the modulation of neurotrophin receptors aimed at formation and regeneration of axons has been proposed as a strategy to delay the progression of neurodegeneration and has gained relevance as new techniques for early diagnosis arise. Based on these premises, we investigated the potential of doxycycline to mimic the effects of Nerve Growth Factor (NGF) with focus on the signaling pathways and neuronal modulators of neurite initiation, growth and branching. We used PC12 cells, a neuronal model widely employed to study the neurotrophic pathways and mechanisms induced by NGF. Results showed that doxycycline induced neurite outgrowth via activation of the trkA receptor and the downstream signaling pathways, PI3K/Akt and MAPK/ERK, without inducing the expression of NGF. Doxycycline also increased the expression of GAP-43, synapsin I and NF200, proteins involved in axonal and synaptic plasticity. Altogether, these data demonstrate, for the first time, the neurotrophic potential of doxycycline, which might be useful to restore the neuronal connectivity lost at the initial phase of neurodegeneration.
Assuntos
Antibacterianos/farmacologia , Doxiciclina/farmacologia , Fator de Crescimento Neural/metabolismo , Animais , Carbazóis/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Proteína GAP-43/metabolismo , Alcaloides Indólicos/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Fator de Crescimento Neural/farmacologia , Proteínas de Neurofilamentos/metabolismo , Crescimento Neuronal/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Células PC12 , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Receptor trkA/antagonistas & inibidores , Receptor trkA/metabolismo , Transdução de Sinais/efeitos dos fármacos , Sinapsinas/metabolismoRESUMO
Protein S-acylation is a reversible post-translational modification involving the addition of fatty acids to cysteines and is catalyzed by transmembrane protein acyltransferases (PATs) mainly expressed at the Golgi complex. In case of soluble proteins, S-acylation confers stable membrane attachment. Myristoylation or farnesylation of many soluble proteins constitutes the initial transient membrane adsorption step prior to S-acylation. However, some S-acylated soluble proteins, such as the neuronal growth-associated protein Growth-associated protein-43 (GAP-43), lack the hydrophobic modifications required for this initial membrane interaction. The signals for GAP-43 S-acylation are confined to the first 13 amino acids, including the S-acylatable cysteines 3 and 4 embedded in a hydrophobic region, followed by a cluster of basic amino acids. We found that mutation of critical basic amino acids drastically reduced membrane interaction and hence S-acylation of GAP-43. Interestingly, acute depletion of phosphatidylinositol 4-phosphate (PtdIns4P) at the Golgi complex reduced GAP-43 membrane binding, highlighting a new, pivotal role for this anionic lipid and supporting the idea that basic amino acid residues are involved in the electrostatic interactions between GAP-43 and membranes of the Golgi complex where they are S-acylated.
Assuntos
Proteína 4 Homóloga a Disks-Large/metabolismo , Proteína GAP-43/metabolismo , Fosfatos de Fosfatidilinositol/metabolismo , Processamento de Proteína Pós-Traducional , Rede trans-Golgi/metabolismo , Acilação , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Células CHO , Sequência Conservada , Cricetulus , Proteína 4 Homóloga a Disks-Large/química , Proteína 4 Homóloga a Disks-Large/genética , Proteína GAP-43/química , Proteína GAP-43/genética , Interações Hidrofóbicas e Hidrofílicas , Eletricidade Estática , Fatores de Tempo , Rede trans-Golgi/genéticaRESUMO
Cisplatin is a highly effective chemotherapeutic drug that is toxic to the peripheral nervous system. Findings suggest that axons are early targets of the neurotoxicity of cisplatin. Although many compounds have been reported as neuroprotective, there is no effective treatment against the neurotoxicity of cisplatin. Caffeic acid phenethyl ester (CAPE) is a propolis component with neuroprotective potential mainly attributed to antioxidant and anti-inflammatory mechanisms. We have recently demonstrated the neurotrophic potential of CAPE in a cellular model of neurotoxicity related to Parkinson's disease. Now, we have assessed the neurotrophic and neuroprotective effects of CAPE against cisplatin-induced neurotoxicity in PC12 cells. CAPE (10 µM) attenuated the inhibition of neuritogenesis and the downregulation of markers of neuroplasticity (GAP-43, synapsin I, synaptophysin, and 200-kD neurofilament) induced by cisplatin (5 µM). This concentration of cisplatin does not affect cell viability, and it was used in order to assess the early neurotoxic events triggered by cisplatin. When a lethal dose of cisplatin was used (IC50 = 32 µM), CAPE (10 µM) increased cell viability. The neurotrophic effect of CAPE is not dependent on NGF nor is it additive to the effect of NGF, but it might involve the activation of the NGF-high-affinity receptors (trkA). The involvement of other neurotrophin receptors such as trkB and trkC is unlikely. This is the first study to demonstrate the protective potential of CAPE against the neurotoxicity of cisplatin and to suggest the involvement of trkA receptors in the neuroprotective mechanism of CAPE. Based on these findings, the beneficial effect of CAPE on cisplatin-induced peripheral neuropathy should be further investigated.
Assuntos
Ácidos Cafeicos/farmacologia , Cisplatino/farmacologia , Fator de Crescimento Neural/metabolismo , Fármacos Neuroprotetores/farmacologia , Neurotoxinas/farmacologia , Álcool Feniletílico/análogos & derivados , Transdução de Sinais/efeitos dos fármacos , Análise de Variância , Animais , Diferenciação Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Proteína GAP-43/metabolismo , Neuroblastoma/patologia , Proteínas de Neurofilamentos/metabolismo , Crescimento Neuronal/efeitos dos fármacos , Células PC12/efeitos dos fármacos , Álcool Feniletílico/farmacologia , Ratos , Sinapsinas/metabolismo , Sinaptofisina/metabolismoRESUMO
Angiotensin-(1-7) (Ang [1-7]) and its receptor Mas are involved in a number of physiological processes, including control of arterial pressure and modulation of nervous system actions. However, the involvement of the Ang-(1-7)/Mas axis in peripheral nerve injury has not been investigated. Using a model of sciatic nerve injury in mice, we demonstrated opposing changes in Mas receptor expression at days 2 and 14 post-injury. Mas receptor expression was more intense 2days after the nerve lesion, compared with the intensity of the intact nerve. At this time point, the sciatic nerve functional index was -20. At day 14 after the lesion, the intensity of the immunostaining labeling in longitudinal sections of the nerve was reduced (â¼30%) and the functional index increased +36 (gait improvement). In the axotomized group treated with A779 (a Mas receptor antagonist), the functional recovery index decreased in relation to the untreated axotomized group. The Mas receptor inhibitor also altered the intensity of labeling of S-100, GAP43, and IBA-1 (morphological features compatible with delayed axon growth). This study demonstrated that Ang-(1-7)/Mas axis activity was differentially modulated in the acute and post-acute stages of nerve injury.
Assuntos
Nervos Periféricos/efeitos dos fármacos , Nervos Periféricos/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Neuropatia Ciática/metabolismo , Angiotensina II/análogos & derivados , Angiotensina II/farmacologia , Animais , Proteína GAP-43/metabolismo , Masculino , Camundongos , Fragmentos de Peptídeos/farmacologia , Nervos Periféricos/patologia , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Proteínas S100/metabolismoRESUMO
The olfactory epithelium (OE) has the remarkable capability to constantly replace olfactory receptor neurons (ORNs) due to the presence of neural stem cells (NSCs). For this reason, the OE provides an excellent model to study neurogenesis and neuronal differentiation. In the present work, we induced neuronal degeneration in the OE of Xenopus laevis larvae by bilateral axotomy of the olfactory nerves. We found that axotomy induces specific- neuronal death through apoptosis between 24 and 48h post-injury. In concordance, there was a progressive decrease of the mature-ORN marker OMP until it was completely absent 72h post-injury. On the other hand, neurogenesis was evident 48h post-injury by an increase in the number of proliferating basal cells as well as NCAM-180- GAP-43+ immature neurons. Mature ORNs were replenished 21 days post-injury and the olfactory function was partially recovered, indicating that new ORNs were integrated into the olfactory bulb glomeruli. Throughout the regenerative process no changes in the expression pattern of the neurotrophin Brain Derivate Neurotrophic Factor were observed. Taken together, this work provides a sequential analysis of the neurodegenerative and subsequent regenerative processes that take place in the OE following axotomy. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 77: 1308-1320, 2017.
Assuntos
Axotomia , Degeneração Neural/etiologia , Degeneração Neural/patologia , Mucosa Olfatória/patologia , Traumatismos do Nervo Olfatório/patologia , Regeneração/fisiologia , Animais , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Caspase 3/metabolismo , Diferenciação Celular/fisiologia , Proliferação de Células , Proteína GAP-43/metabolismo , Regulação da Expressão Gênica/fisiologia , Queratina-2/metabolismo , Moléculas de Adesão de Célula Nervosa/metabolismo , Proteína de Marcador Olfatório/metabolismo , Traumatismos do Nervo Olfatório/etiologia , Recuperação de Função Fisiológica/fisiologia , Olfato/fisiologia , Fatores de Tempo , Xenopus laevisRESUMO
During postnatal development, neural circuits are extremely dynamic and develop precise connection patterns that emerge as a result of the elimination of synaptic terminals, a process instructed by molecular cues and patterns of electrical activity. In the rodent visual system, this process begins during the first postnatal week and proceeds during the second and third postnatal weeks as spontaneous retinal activity and finally use-dependent fine tuning takes place. Reelin is a large extracellular matrix glycoprotein able to affect several steps of brain development, from neuronal migration to the maturation of dendritic spines and use-dependent synaptic development. In the present study, we investigated the role of reelin on the topographical refinement of primary sensory connections studying the development of retinal ganglion cell axon terminals in the rat superior colliculus. We found that reelin levels in the visual layers of the superior colliculus are the highest between the second and third postnatal weeks. Blocking reelin signaling with a neutralizing antibody (CR-50) from PND 7 to PND 14 induced a non-specific sprouting of ipsilateral retinocollicular axons outside their typical distribution of discrete patches of axon terminals. Also we found that reelin blockade resulted in reduced levels of phospho-GAP43, increased GluN1 and GluN2B-NMDA subunits and decreased levels of GAD65 content in the visual layers of the superior colliculus. The results suggest that reelin signaling is associated with the maturation of excitatory and inhibitory synaptic machinery influencing the development and fine tuning of topographically organized neural circuits during postnatal development.
Assuntos
Moléculas de Adesão Celular Neuronais/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Células Ganglionares da Retina/metabolismo , Serina Endopeptidases/metabolismo , Colículos Superiores/crescimento & desenvolvimento , Colículos Superiores/metabolismo , Animais , Animais Recém-Nascidos , Western Blotting , Moléculas de Adesão Celular Neuronais/antagonistas & inibidores , Proteínas da Matriz Extracelular/antagonistas & inibidores , Proteína GAP-43/metabolismo , Glutamato Descarboxilase/metabolismo , Imuno-Histoquímica , Proteínas do Tecido Nervoso/antagonistas & inibidores , Técnicas de Rastreamento Neuroanatômico , Fosforilação/fisiologia , Ratos , Receptores de N-Metil-D-Aspartato/metabolismo , Proteína Reelina , Células Ganglionares da Retina/citologia , Colículos Superiores/citologia , Vias Visuais/citologia , Vias Visuais/crescimento & desenvolvimento , Vias Visuais/metabolismoRESUMO
The aim of this study was to investigate whether exogenous retinoic acid (RA) can upregulate the mRNA and protein expression of growth-associated protein 43 (GAP-43), thereby promoting brain functional recovery in a rat distal middle cerebral artery occlusion (MCAO) model of ischemia. A total of 216 male Sprague Dawley rats weighing 300-320 g were divided into 3 groups: sham-operated group, MCAO+vehicle group and MCAO+RA group. Focal cortical infarction was induced with a distal MCAO model. The expression of GAP-43 mRNA and protein in the ipsilateral perifocal region was assessed using qPCR and immunocytochemistry at 1, 3, 7, 14, 21, and 28 days after distal MCAO. In addition, an intraperitoneal injection of RA was given 12 h before MCAO and continued every day until the animal was sacrificed. Following ischemia, the expression of GAP-43 first increased considerably and then decreased. Administration of RA reduced infarction volume, promoted neurological functional recovery and upregulated expression of GAP-43. Administration of RA can ameliorate neuronal damage and promote nerve regeneration by upregulating the expression of GAP-43 in the perifocal region after distal MCAO.
Assuntos
Proteína GAP-43/metabolismo , Expressão Gênica/efeitos dos fármacos , Infarto da Artéria Cerebral Média/prevenção & controle , Fármacos Neuroprotetores/farmacologia , Tretinoína/farmacologia , Regulação para Cima/efeitos dos fármacos , Animais , Isquemia Encefálica/prevenção & controle , Proteína GAP-43/genética , Imuno-Histoquímica , Infarto da Artéria Cerebral Média/metabolismo , Infarto da Artéria Cerebral Média/patologia , Masculino , Distribuição Aleatória , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Reprodutibilidade dos Testes , Fatores de TempoRESUMO
BACKGROUND: Immunization with neural derived peptides (INDP) as well as scar removal-separately-have shown to induce morphological and functional improvement after spinal cord injury (SCI). In the present study, we compared the effect of INDP alone versus INDP with scar removal on motor recovery, regeneration-associated and cytokine gene expression, and axonal regeneration after chronic SCI. Scar removal was conducted through a single incision with a double-bladed scalpel along the stump, and scar renewal was halted by adding α,α'-dipyridyl. RESULTS: During the chronic injury stage, two experiments were undertaken. The first experiment was aimed at testing the therapeutic effect of INDP combined with scar removal. Sixty days after therapeutic intervention, the expression of genes encoding for TNFα, IFNγ, IL4, TGFß, BDNF, IGF1, and GAP43 was evaluated at the site of injury. Tyrosine hydroxylase and 5-hydroxytryptamine positive fibers were also studied. Locomotor evaluations showed a significant recovery in the group treated with scar removal + INDP. Moreover; this group presented a significant increase in IL4, TGFß, BDNF, IGF1, and GAP43 expression, but a decrease of TNFα and IFNγ. Also, the spinal cord of animals receiving both treatments presented a significant increase of serotonergic and catecholaminergic fibers as compared to other the groups. The second experiment compared the results of the combined approach versus INDP alone. Rats receiving INDP likewise showed improved motor recovery, although on a lesser scale than those who received the combined treatment. An increase in inflammation and regeneration-associated gene expression, as well as in the percentage of serotonergic and catecholaminergic fibers was observed in INDP-treated rats to a lesser degree than those in the combined therapy group. CONCLUSIONS: These findings suggest that INDP, both alone and in combination with scar removal, could modify the non-permissive microenvironment prevailing at the chronic phase of SCI, providing the opportunity of improving motor recovery.
Assuntos
Cicatriz/metabolismo , Locomoção/efeitos dos fármacos , Neuropeptídeos/administração & dosagem , Traumatismos da Medula Espinal/imunologia , Traumatismos da Medula Espinal/metabolismo , Vacinação , Animais , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Doença Crônica , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Proteína GAP-43/metabolismo , Fator de Crescimento Insulin-Like I/metabolismo , Interferon gama/metabolismo , Interleucina-4/metabolismo , Neuropeptídeos/uso terapêutico , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica , Traumatismos da Medula Espinal/tratamento farmacológico , Regeneração da Medula Espinal/genética , Fator de Crescimento Transformador beta/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Some organophosphorus compounds (OPs) induce a neurodegenerative disorder known as organophosphate-induced delayed neuropathy (OPIDN), which is related to irreversible inhibition of neuropathy target esterase (NTE) and impairment of neurite outgrowth. The present study addresses the effects of trichlorfon, mipafox (neuropathic model) and paraoxon (non-neuropathic model) on neurite outgrowth and neuroplasticity-related proteins in retinoic-acid-stimulated SH-SY5Y cells, a cellular model widely used to study the neurotoxicity of OPs. Mipafox (20µM) decreased cellular differentiation and the expression of neurofilament 200 (NF-200), growth associated- (GAP-43) and synaptic proteins (synapsin I and synaptophysin); whereas paraoxon (300µM) induced no effect on cellular differentiation, but significant decrease of NF-200, GAP-43, synapsin I and synaptophysin as compared to controls. However, the effects of paraoxon on these proteins were significantly lower than the effects of mipafox. In conclusion, axonal cytoskeletal proteins, as well as axonal plasticity-related proteins are more effectively affected by neuropathic (mipafox) than by non-neuropathic (paraoxon) OPs, suggesting that they might play a role in the mechanism of OPIDN. At high concentration (1mM), trichlorfon induced effects similar to those of the neuropathic OP, mipafox (20µM), but also caused high inhibition of AChE. Therefore, these effects are unlikely to occur in humans at non-lethal doses of trichlorfon.
Assuntos
Axônios/efeitos dos fármacos , Inibidores da Colinesterase/toxicidade , Inseticidas/toxicidade , Isoflurofato/análogos & derivados , Paraoxon/toxicidade , Triclorfon/toxicidade , Acetilcolinesterase/metabolismo , Hidrolases de Éster Carboxílico/metabolismo , Linhagem Celular Tumoral , Citoesqueleto/efeitos dos fármacos , Proteína GAP-43/metabolismo , Humanos , Isoflurofato/toxicidade , L-Lactato Desidrogenase/metabolismo , Plasticidade Neuronal , Sinapsinas/metabolismoRESUMO
The aim of this study was to investigate whether exogenous retinoic acid (RA) can upregulate the mRNA and protein expression of growth-associated protein 43 (GAP-43), thereby promoting brain functional recovery in a rat distal middle cerebral artery occlusion (MCAO) model of ischemia. A total of 216 male Sprague Dawley rats weighing 300–320 g were divided into 3 groups: sham-operated group, MCAO+vehicle group and MCAO+RA group. Focal cortical infarction was induced with a distal MCAO model. The expression of GAP-43 mRNA and protein in the ipsilateral perifocal region was assessed using qPCR and immunocytochemistry at 1, 3, 7, 14, 21, and 28 days after distal MCAO. In addition, an intraperitoneal injection of RA was given 12 h before MCAO and continued every day until the animal was sacrificed. Following ischemia, the expression of GAP-43 first increased considerably and then decreased. Administration of RA reduced infarction volume, promoted neurological functional recovery and upregulated expression of GAP-43. Administration of RA can ameliorate neuronal damage and promote nerve regeneration by upregulating the expression of GAP-43 in the perifocal region after distal MCAO.
Assuntos
Animais , Masculino , Proteína GAP-43/metabolismo , Expressão Gênica/efeitos dos fármacos , Infarto da Artéria Cerebral Média/prevenção & controle , Fármacos Neuroprotetores/farmacologia , Tretinoína/farmacologia , Regulação para Cima/efeitos dos fármacos , Isquemia Encefálica/prevenção & controle , Proteína GAP-43/genética , Imuno-Histoquímica , Infarto da Artéria Cerebral Média/metabolismo , Infarto da Artéria Cerebral Média/patologia , Distribuição Aleatória , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Reprodutibilidade dos Testes , Fatores de TempoRESUMO
Cisplatin is the most effective and neurotoxic platinum chemotherapeutic agent. It induces a peripheral neuropathy characterized by distal axonal degeneration that might progress to degeneration of cell bodies and apoptosis. Most symptoms occur nearby distal axonal branches and axonal degeneration might induce peripheral neuropathy regardless neuronal apoptosis. The toxic mechanism of cisplatin has been mainly associated with DNA damage, but cisplatin might also affect neurite outgrowth. Nevertheless, the neurotoxic mechanism of cisplatin remains unclear. We investigated the early effects of cisplatin on axonal plasticity by using non-cytotoxic concentrations of cisplatin and PC12 cells as a model of neurite outgrowth and differentiation. PC12 cells express NGF-receptors (trkA) and respond to NGF by forming neurites, branches and synaptic vesicles. For comparison, we used a neuronal model (SH-SY5Y cells) that does not express trkA nor responds to NGF. Cisplatin did not change NGF expression in PC12 cells and decreased neurite outgrowth in both models, suggesting a NGF/trkA independent mechanism. It also reduced axonal growth (GAP-43) and synaptic (synapsin I and synaptophysin) proteins in PC12 cells, without inducing mitochondrial damage or apoptosis. Therefore, cisplatin might affect axonal plasticity before DNA damage, NGF/trkA down-regulation, mitochondrial damage or neuronal apoptosis. This is the first study to show that neuroplasticity-related proteins might be early targets of the neurotoxic action of cisplatin and their role on cisplatin-induced peripheral neuropathy should be investigated in vivo.
Assuntos
Cisplatino/farmacologia , Fator de Crescimento Neural/metabolismo , Crescimento Neuronal/efeitos dos fármacos , Plasticidade Neuronal/efeitos dos fármacos , Animais , Axônios/efeitos dos fármacos , Axônios/metabolismo , Diferenciação Celular/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Proteína GAP-43/metabolismo , Neuritos/efeitos dos fármacos , Neuritos/fisiologia , Células PC12 , Ratos , Receptores de Fator de Crescimento Neural/metabolismoRESUMO
The aim of this study was to explore the immune protective mechanism of rMOMP protein vaccine in intraocular hypertension and retinal optic nerve injury in rats. The rMOMP protein ophthalmic vaccine was prepared and quality-controlled. Sixty normal adult SD rats were randomly divided into two groups to establish a chronic ocular hypertension model and an optic nerve injury model. The model rats were vaccinated with rMOMP-CS ophthalmic vaccine. Fluorogold retrograde tracing was used to observe retinal ganglion cells, and an immunofluorescence method to determine the expression of retinal GAP43, CD3, BDNF, and GDNF. rMOMP protein ophthalmic vaccine met the requirements for medicinal use. The number of retinal ganglion cells (RGCs) of the rMOMP-CS group in the chronic ocular hypertension model was significantly higher than that of the CS group (P < 0.05). The count of RGCs of the rMOMP-CS group in the optic nerve clamping injury model was significantly higher than that of the CS group (P < 0.01). Thus, rMOMP protein ophthalmic vaccine can induce an increase in the expression of retinal neurotrophic factors, thereby exerting a protective effect on damaged retinal optic nerve.
Assuntos
Proteínas da Membrana Bacteriana Externa/imunologia , Sistema Imunitário/imunologia , Hipertensão Ocular/imunologia , Traumatismos do Nervo Óptico/imunologia , Vacinas/imunologia , Animais , Proteínas da Membrana Bacteriana Externa/genética , Fator Neurotrófico Derivado do Encéfalo/imunologia , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Complexo CD3/imunologia , Complexo CD3/metabolismo , Doença Crônica , Feminino , Proteína GAP-43/imunologia , Proteína GAP-43/metabolismo , Fator Neurotrófico Derivado de Linhagem de Célula Glial/imunologia , Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Sistema Imunitário/efeitos dos fármacos , Masculino , Microscopia Confocal , Hipertensão Ocular/metabolismo , Hipertensão Ocular/prevenção & controle , Traumatismos do Nervo Óptico/metabolismo , Traumatismos do Nervo Óptico/prevenção & controle , Substâncias Protetoras/administração & dosagem , Distribuição Aleatória , Ratos Sprague-Dawley , Proteínas Recombinantes/imunologia , Retina/imunologia , Retina/metabolismo , Células Ganglionares da Retina/imunologia , Células Ganglionares da Retina/metabolismo , Estilbamidinas/metabolismo , Vacinação/métodos , Vacinas/administração & dosagemRESUMO
Growth-associated protein 43 (GAP-43) is a neuronal phosphoprotein associated with initial axonal outgrowth and synaptic remodeling and recent work also suggests its involvement in cell cycle control. The complex expression of GAP-43 features transcriptional and posttranscriptional components. However, in some conditions, GAP-43 gene expression is controlled primarily by the interaction of stabilizing or destabilizing RNA-binding proteins (RBPs) with adenine and uridine (AU)-rich instability elements (AREs) in its 3'UTR. Like GAP-43, many proteins involved in cell proliferation are encoded by ARE-containing mRNAs, some of which codify cell-cycle-regulating proteins including cyclin D1. Considering that GAP-43 and cyclin D1 mRNA stabilization may depend on similar RBPs, this study evaluated the participation of GAP-43 in cell cycle control and its underlying mechanisms, particularly the possible role of its 3'UTR, using GAP-43-transfected NIH-3T3 fibroblasts. Our results show an arrest in cell cycle progression in the G0/G1 phase. This arrest may be mediated by the competition of GAP-43 3'UTR with cyclin D1 3'UTR for the binding of Hu proteins such as HuR, which may lead to a decrease in cyclin D1 expression. These results might lead to therapeutic applications involving the use of sequences in the B region of GAP-43 3'UTR to slow down cell cycle progression.
Assuntos
Proteína GAP-43/metabolismo , Regiões 3' não Traduzidas , Animais , Adesão Celular , Ciclo Celular , Proliferação de Células , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Quinases Ciclina-Dependentes/metabolismo , Ciclinas/metabolismo , Ativação Enzimática , Proteína GAP-43/genética , Camundongos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Células NIH 3T3 , Proteínas de Ligação a RNA/metabolismoRESUMO
OBJECTIVES: We, herein, analyzed the effect of swimming on nociception threshold and peripheral nerve regeneration in lean and obese rats submitted to median nerve compression. METHODS: To induce obesity, newborn male Wistar rats received injections of monosodium glutamate (MSG), whereas the control (CTL) group received saline. The animals were separated into 6 groups; control and obese (CTL and MSG), control and obese with lesion (CTL LES and MSG LES), and control and obese with lesion submitted to physical exercise (CTL LES PE and MSG LES PE). RESULTS: Median nerve compression reduced nociception threshold in CTL LES and MSG LES rats. Swimming effectively altered nociception only in CTL LES PE animals. Lean and obese animals displayed histological differences, when compared to sedentary animals, and exercise improved axon regeneration in both groups. The brain-derived neurotrophic factor and GAP 43 protein expression was greater in animals submitted to nervous compression without alteration by exercise. DISCUSSION: In conclusion, swimming, a conservative treatment for peripheral nerve lesions, was not able to improve the nociception threshold in obese rats.
Assuntos
Síndromes de Compressão Nervosa/fisiopatologia , Síndromes de Compressão Nervosa/reabilitação , Nociceptividade/fisiologia , Obesidade/complicações , Limiar da Dor/fisiologia , Natação , Análise de Variância , Animais , Animais Recém-Nascidos , Índice de Massa Corporal , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Modelos Animais de Doenças , Proteína GAP-43/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Masculino , Nervo Mediano/fisiopatologia , Síndromes de Compressão Nervosa/patologia , Obesidade/induzido quimicamente , Medição da Dor , Condicionamento Físico Animal , Ratos , Ratos Wistar , Glutamato de Sódio/farmacologiaRESUMO
The vomeronasal organ (VNO), also known as the Jacobson's organ, is a bilateral chemosensory organ found at the base of the nasal cavity specialized for the detection of higher-molecular weight (non-volatile) chemostimuli. It has been linked to pheromone detection. The VNO has been well studied in nocturnal lemurs and lorises, but poorly studied in diurnal/cathemeral species despite the large repertoire of olfactory behaviors noted in species such as Lemur catta. Here, the VNO and associated structures were studied microanatomically in one adult female and one adult male L. catta. Traditional and immunohistochemical procedures demonstrate the VNO epithelium consists of multiple rows of sensory neurons. Immunoreactivity to Growth-associated protein 43 (GAP43) indicates the VNO is postnatally neurogenic. In volume, the VNO neuroepithelium scales similarly to palatal length compared to nocturnal strepsirrhines. Numerous taste buds present at the oral opening to the nasopalatine duct, with which the VNO communicates, provide an additional (or alternative) explanation for the flehmen behavior that has been observed in this species. The VNO of L. catta is shown to be microanatomically comparable to that of nocturnal strepsirrhines. Like nocturnal strepsirrhines, the VNO of L. catta may be functional in the reception of high-molecular weight secretions.
Assuntos
Lemur/anatomia & histologia , Órgão Vomeronasal/anatomia & histologia , Animais , Feminino , Proteína GAP-43 , Imuno-Histoquímica , Lemur/fisiologia , Masculino , Neurônios Receptores Olfatórios/citologia , Papilas Gustativas/anatomia & histologia , Órgão Vomeronasal/fisiologiaRESUMO
Epidermal neural crest stem cells (EPI-NCSCs), which reside in the bulge of hair follicles, are attractive candidates for several applications in cell therapy, drug screening and tissue engineering. As suggested remnants of the embryonic neural crest (NC) in an adult location, EPI-NCSCs are able to generate a wide variety of cell types and are readily accessible by a minimally invasive procedure. Since the combination of epidermal growth factor (EGF) and fibroblast growth factor type 2 (FGF2) is mitogenic and promotes the neuronal commitment of various stem cell populations, we examined its effects in the proliferation and neuronal potential of mouse EPI-NCSCs. By using a recognized culture protocol of bulge whiskers follicles, we were able to isolate a population of EPI-NCSCs, characterized by the migratory potential, cell morphology and expression of phenotypic markers of NC cells. EPI-NCSCs expressed neuronal, glial and smooth muscle markers and exhibited the NC-like fibroblastic morphology. The treatment with the combination EGF and FGF2, however, increased their proliferation rate and promoted the acquisition of a neuronal-like morphology accompanied by reorganization of neural cytoskeletal proteins ßIII-tubulin and nestin, as well as upregulation of the pan neuronal marker ßIII-tubulin and down regulation of the undifferentiated NC, glial and smooth muscle cell markers. Moreover, the treatment enhanced the response of EPI-NCSCs to neurogenic stimulation, as evidenced by induction of GAP43, and increased expression of Mash-1 in neuron-like cell, both neuronal-specific proteins. Together, the results suggest that the combination of EGF-FGF2 stimulates the proliferation and improves the neuronal potential of EPI-NCSCs similarly to embryonic NC cells, ES cells and neural progenitor/stem cells of the central nervous system and highlights the advantage of using EGF-FGF2 in neuronal differentiation protocols.
Assuntos
Fator de Crescimento Epidérmico/metabolismo , Epiderme/metabolismo , Fator 2 de Crescimento de Fibroblastos/metabolismo , Crista Neural/metabolismo , Células-Tronco Neurais/metabolismo , Neurônios/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Biomarcadores/metabolismo , Diferenciação Celular/fisiologia , Proliferação de Células , Regulação para Baixo/fisiologia , Células Epiteliais/metabolismo , Proteína GAP-43/metabolismo , Folículo Piloso/metabolismo , Camundongos , Células-Tronco Multipotentes/metabolismo , Miócitos de Músculo Liso/metabolismo , Regulação para Cima/fisiologiaRESUMO
Reactionary dentin formation is an adaptive secretory response mediated by odontoblasts to moderate dentin injury. The implications of this process for neuroimmune interactions operating to contain pathogens have not been fully appreciated. The purpose of the present study was to describe the relationship between reactionary dentinogenesis, the neurogenic changes of dental pulp innervation, and dendritic cell recruitment to caries progression, using a comparative immunohistochemical approach in human teeth from young adult individuals. Reactionary dentin formation during dentin caries progression is associated with changes in the integrity of junctional complexes within the odontoblast layer. Diminished coexpression of Cx43 and zonula occludens 1 implies a reduced level of intercellular connectivity between odontoblasts. Dentin caries also causes overexpression of growth-associated protein 43, a modulator of neural plasticity that promotes extensive sprouting of nerve endings into the reactionary dentin matrix. At the same time, an elevated number of HLA-DR-positive dendritic cells infiltrate the odontoblast layer and subsequently invade reactionary dentin formed underneath the early caries-affected regions. Simultaneous odontoblast layer remodeling, nerve fiber sprouting, and activation of dendritic cells during caries progression suggest a coordinated neuroimmune response to fight caries pathogen invasion and to promote dentin-pulp healing. We propose that reactionary dentin formation hinders pathogen invasion and supports defensive neuroimmune interactions against infection. The eventual understanding of this complex scenario may contribute to the development of novel approaches to dental caries treatment.