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1.
Nat Neurosci ; 22(11): 1913-1924, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31591560

RESUMO

Axonal injury results in regenerative success or failure, depending on whether the axon lies in the peripheral or the CNS, respectively. The present study addresses whether epigenetic signatures in dorsal root ganglia discriminate between regenerative and non-regenerative axonal injury. Chromatin immunoprecipitation for the histone 3 (H3) post-translational modifications H3K9ac, H3K27ac and H3K27me3; an assay for transposase-accessible chromatin; and RNA sequencing were performed in dorsal root ganglia after sciatic nerve or dorsal column axotomy. Distinct histone acetylation and chromatin accessibility signatures correlated with gene expression after peripheral, but not central, axonal injury. DNA-footprinting analyses revealed new transcriptional regulators associated with regenerative ability. Machine-learning algorithms inferred the direction of most of the gene expression changes. Neuronal conditional deletion of the chromatin remodeler CCCTC-binding factor impaired nerve regeneration, implicating chromatin organization in the regenerative competence. Altogether, the present study offers the first epigenomic map providing insight into the transcriptional response to injury and the differential regenerative ability of sensory neurons.


Assuntos
Axônios/fisiologia , Epigenômica , Gânglios Espinais/fisiologia , Regeneração Nervosa/fisiologia , Células Receptoras Sensoriais/fisiologia , Acetilação , Algoritmos , Animais , Fator de Ligação a CCCTC/genética , Cromatina/metabolismo , Feminino , Gânglios Espinais/lesões , Expressão Gênica , Histonas/metabolismo , Aprendizado de Máquina , Masculino , Camundongos , Camundongos Transgênicos , Nervo Isquiático/lesões , Análise de Sequência de RNA
2.
Neurochem Res ; 44(5): 1214-1227, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30859436

RESUMO

Neuropathic pain (NP) has complicated pathogenesis as it mainly involves a lesion or dysfunction of the somatosensory nervous system and its clinical treatment remains challenging. Chronic constriction injury (CCI) model is a widely used neuropathic pain model and involved in mechanisms including both nerve inflammatory and injury. Cytokines and their receptors play essential roles in the occurrence and persistence of neuropathic pain, but the underlying mechanisms have not well been understood. Therefore, Interleukin-1 receptor-associated kinase 1 (IRAK1) is chosen to explore the possible mechanisms of NP. In the present study, IRAK1 was found to persistently increase in the dorsal root ganglion (DRG) and spinal cord (SC) during CCI detected by western blot. The staining further confirmed that IRAK1 was mainly co-located in the DRG astrocytes or SC neurons, but less in the DRG microglia or SC astrocytes. Moreover, the region of increased IRAK1 expression was observed in superficial laminae of the spinal dorsal horn, which was the nociceptive neuronal expression domain, suggesting that IRAK1 may mediated CCI-induced pain by nociceptive primary afferent. In addition, intrathecal injection of Toll-like receptor 4 (TLR4) inhibitor or IRAK1 siRNA decreased the expression of IRAK1 accompanied with the alleviation of CCI-induced neuropathic pain. The upregulation of p-NF-κB expression was reversed by IRAK1 siRNA in SC, and intrathecal injection of p-NF-κB inhibitor relieved neuropathic pain. Taking together, targeting IRAK1 may be a potential treatment for chronic neuropathic pain.


Assuntos
Gânglios Espinais/metabolismo , Neuralgia/metabolismo , Neuralgia/fisiopatologia , Nervo Isquiático/lesões , Animais , Doença Crônica , Constrição , Gânglios Espinais/lesões , Hiperalgesia/metabolismo , Masculino , Microglia/metabolismo , Nociceptores/metabolismo , Ratos Sprague-Dawley , Receptores de Interleucina-1/metabolismo , Medula Espinal/metabolismo , Medula Espinal/fisiopatologia
3.
World Neurosurg ; 125: e1050-e1056, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30776514

RESUMO

OBJECTIVE: To retrospectively evaluate long-term efficacy and safety of dorsal root entry zone (DREZ) lesion for treatment of neuropathic pain within the lower extremities and perineal region after thoracolumbar spine fracture. METHODS: Forty-two patients were treated with posterior laminectomy under general anesthesia. The DREZ regions of the spinal cord segments were ablated under a microscope. Data regarding pain relief, pain variation over time, and postoperative complications were collected. The relationship between injured spinal column segment, spinal cord, nerve root, and pain territory were analyzed retrospectively. RESULTS: Spinal column injury segments were located between T12 and L4. Pain territories were distributed between the T11 and S5 dermatomes with varying ranges, at an average of 2-6 segments higher than the spinal cord injury segments. Pain relief rate was 100% in 21 patients (50.0%) and was over 50% in 14 patients (33.3%). Eighteen patients (42.9%) developed temporary tingling in the upper edge of the spinal cord lesion segment after surgery. Of the 4 patients with unilateral lower extremity pain, 2 developed postoperative persistent pain in the contralateral lower extremity. CONCLUSIONS: For patients with neuropathic pain of the lower extremities and/or the perineal region after thoracolumbar spine fracture, pain within the lower extremities was mostly because of nerve root injury. Pain in the perineal region caused by L1 fracture was attributed to spinal cord injury segmental pain. Nerve root injury pain had a good prognosis after DREZ lesion; the effect of DREZ lesion for spinal cord injury segmental pain may be uncertain.


Assuntos
Gânglios Espinais/cirurgia , Vértebras Lombares/lesões , Neuralgia/cirurgia , Fraturas da Coluna Vertebral/complicações , Vértebras Torácicas/lesões , Feminino , Gânglios Espinais/lesões , Humanos , Masculino , Neuralgia/etiologia , Procedimentos Neurocirúrgicos/métodos , Medição da Dor , Estudos Retrospectivos , Resultado do Tratamento
4.
Neurol India ; 67(Supplement): S32-S37, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30688230

RESUMO

Peripheral nerve and brachial plexus injuries typically cause severe impairment in the affected limb. The incidence of neuropathic pain is high, reaching up to 95% of cases, especially if cervical root avulsion has occurred. Neuropathic pain results from damage to the somatosensory system, and its progression towards chronicity depends upon disruptions affecting both the peripheral and central nervous system. Managing these painful conditions is complex and must be accomplished by a multidisciplinary team, starting with first-line pharmacological therapies like tricyclic antidepressants and calcium channel ligands, combined physical and occupational therapy, transcutaneous electrical stimulation and psychological support. For patients refractory to the initial measures, several neurosurgical options are available, including nerve decompression or reconstruction and ablative/modulatory procedures.


Assuntos
Plexo Braquial/lesões , Neuralgia/terapia , Traumatismos dos Nervos Periféricos/complicações , Plexo Braquial/fisiopatologia , Gânglios Espinais/lesões , Gânglios Espinais/fisiopatologia , Humanos , Neuralgia/etiologia , Neuralgia/fisiopatologia , Traumatismos dos Nervos Periféricos/fisiopatologia , Resultado do Tratamento
5.
Neuroscience ; 402: 51-65, 2019 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-30699332

RESUMO

Low back pain is a common cause of chronic pain and disability. It is modeled in rodents by chronically compressing the lumbar dorsal root ganglia (DRG) with small metal rods, resulting in ipsilateral mechanical and cold hypersensitivity, and hyperexcitability of sensory neurons. Sodium channels are implicated in this hyperexcitability, but the responsible isoforms are unknown. In this study, we used siRNA-mediated knockdown of the pore-forming NaV1.6 and regulatory NaVß4 sodium channel isoforms that have been previously implicated in a different model of low back pain caused by locally inflaming the L5 DRG. Knockdown of either subunit markedly reduced spontaneous pain and mechanical and cold hypersensitivity induced by DRG compression, and reduced spontaneous activity and hyperexcitability of sensory neurons with action potentials <1.5 msec (predominately cells with myelinated axons, based on conduction velocities measured in a subset of cells) 4 days after DRG compression. These results were similar to those previously obtained in the DRG inflammation model and some neuropathic pain models, in which sensory neurons other than nociceptors seem to play key roles. The cytokine profiles induced by DRG compression and DRG inflammation were also very similar, with upregulation of several type 1 pro-inflammatory cytokines and downregulation of type 2 anti-inflammatory cytokines. Surprisingly, the cytokine profile was largely unaffected by NaVß4 knockdown in either model. The NaV1.6 channel, and the NaVß4 subunit that can regulate NaV1.6 to enhance repetitive firing, play key roles in both models of low back pain; targeting the abnormal spontaneous activity they generate may have therapeutic value.


Assuntos
Gânglios Espinais/metabolismo , Dor Lombar/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.6/metabolismo , Neurônios/fisiologia , Subunidade beta-4 do Canal de Sódio Disparado por Voltagem/metabolismo , Potenciais de Ação , Animais , Feminino , Gânglios Espinais/lesões , Gânglios Espinais/fisiopatologia , Inflamação/metabolismo , Mediadores da Inflamação/metabolismo , Dor Lombar/fisiopatologia , Masculino , Modelos Animais , Limiar da Dor , Ratos Sprague-Dawley , Compressão da Medula Espinal , Regulação para Cima
6.
Neurol Res ; 41(3): 265-274, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30570422

RESUMO

OBJECTIVE: Peripheral nerve injury (PNI) is an important global health problem. Nerve Growth Factor (NGF) plays crucial role in the survival, growth, and maintenance of various neurons in the mammalian nervous system, human included. Hericium erinaceus (HE), an edible and medicinal mushroom, has been extensively studied for its neuroprotective properties. In this study, the neuroprotective and neurotogenic effects of HE and NGF were compared on mouse PNI model by using a laser microdissection technique. METHODS: Neuronal cultures were prepared from dorsal root ganglia (DRG) of 6-8 week aged mice, pretreated them with phosphate-buffered saline (PBS), NGF, HE, or the combination of NGF and HE. To model axonal injury in vitro, axons were cut (axotomy) with a microscope-controlled laser beam. Axotomized neurons were imaged under the microscope. Axotomized neurons' survival ratios were calculated using the propidium iodide (PI), which is a red-fluorescent nuclear dye. Their axon lengths were measured using the AxioVision 4.8 software. RESULTS: Although both HE and NGF have neuroprotective and regenerative effects on axotomized peripheral sensory neurons, HE exhibits a higher neuroprotective activity compared to the NGF. The combination of HE and NGF maximizes axonal regeneration ability of axotomized neurons. CONCLUSION: HE has capabilities of preventing the death of neurons and regenerating their axons in the experimental axonal injury model. Our findings provide experimental evidence that HE may serve as a neuroprotective and regenerative candidate for treating peripheral nerve injuries. Present study warrants further investigation of HE as a potential natural compound to remedy PNI.


Assuntos
Agaricales , Axônios/efeitos dos fármacos , Regeneração Nervosa/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Extratos Vegetais/farmacologia , Animais , Axônios/patologia , Axônios/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/lesões , Gânglios Espinais/patologia , Gânglios Espinais/fisiopatologia , Masculino , Camundongos Endogâmicos BALB C , Microdissecção , Fator de Crescimento Neural/farmacologia , Regeneração Nervosa/fisiologia , Traumatismos dos Nervos Periféricos/tratamento farmacológico , Traumatismos dos Nervos Periféricos/patologia , Traumatismos dos Nervos Periféricos/fisiopatologia , Fitoterapia , Distribuição Aleatória
7.
J Ethnopharmacol ; 233: 131-140, 2019 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-30590196

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Tabebuia aurea (Silva Manso) Benth. & Hook. f. ex S. Moore is used as anti-inflammatory, analgesic and antiophidic in traditional medicine, though its pharmacological proprieties are still underexplored. In the bothropic envenoming, pain is a key symptom drove by an intense local inflammatory and neurotoxic event. The antivenom serum therapy is still the main treatment despite its poor local effects against pain and tissue injury. Furthermore, it is limited to ambulatorial niches, giving space for the search of new and more inclusive pharmacological approaches. AIM OF THE STUDY: evaluation of Tabebuia aurea hydroethanolic extract (HEETa) in hyperalgesia and neuronal injury induced by Bothrops mattogrossensis venom (VBm). MATERIALS AND METHODS: Stem barks from Tabebuia aurea were extracted with ethanol and water (7:3, v/v) to yield the extract HEETa. Then, HEETa was analyzed by LC-DAD-MS and its constituents were identified. Snake venoms were extracted from adult specimens of Bothrops mattogrossensis, lyophilized and kept at -20 °C until use. Male Swiss mice, weighting 20-25 g, were used to hyperalgesia (electronic von Frey), motor impairment (Rotarod test) and tissue injury evaluation (histopatology and ATF-3 immunohistochemistry). Therefore, three experimental groups were formed: VBm (1 pg, 1 ng, 0.3 µg, 1 µg, 3 and 6 µg/paw), HEETa orally (180, 540, 720, 810 or 1080 mg/kg; 10 mL/kg, 30 min prior VBm inoculation) and VBm neutralized (VBm: HEETa, 1:100 parts, respectively). In all set of experiments a control (saline group) was used. First, we made a dose-time-response course curve of VBm's induced hyperalgesia. Next, VBm maximum hyperalgesic dose was employed to perform HEETa orally dose-time-response course curve and analyses of VBm neutralized. Paw tissues for histopathology and DRGs were collected from animals inoculated with VBm maximum dose and treated with HEETa antihyperalgesic effective dose or neutralized VBm. Paws were extract two or 72 h after VBm inoculation and DRGs, in the maximum expected time expression of ATF-3 (72 h). RESULTS: From HEETa extract, glycosylated iridoids were identified, such as catalpol, minecoside, verminoside and specioside. VBm induced a time and dose dependent hyperalgesia with its highest effect seen with 3 µg/paw, 2 h after venom inoculation. HEETa effective dose (720 mg/kg) decreased significantly VBm induced hyperalgesia (3 µg/paw) with no motor impairment and signs of acute toxicity. HEETa antihyperalgesic action starts 1.5 h after VBm inoculation and lasted up until 2 h after VBm. Hyperalgesia wasn't reduced by VBm: HEETa neutralization. Histopathology revealed a large hemorragic field 2 h after VBm inoculation and an intense inflammatory infiltrate of polymorphonuclear cells at 72 h. Both HEETa orally and VBm: HEETa groups had a reduced inflammation at 72 h after VBm. Also, the venom significantly induced ATF-3 expression (35.37 ±â€¯3.25%) compared with saline group (4.18 ±â€¯0.68%) which was reduced in HEETa orally (25.87 ±â€¯2.57%) and VBm: HEETa (19.84 ±â€¯2.15%) groups. CONCLUSION: HEETa reduced the hyperalgesia and neuronal injury induced by VBm. These effects could be related to iridoid glycosides detected in HEETa and their intrinsic reported mechanism.


Assuntos
Analgésicos/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Bothrops , Hiperalgesia/tratamento farmacológico , Extratos Vegetais/uso terapêutico , Venenos de Serpentes/toxicidade , Tabebuia , Fator 3 Ativador da Transcrição/metabolismo , Analgésicos/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Gânglios Espinais/lesões , Hiperalgesia/metabolismo , Masculino , Camundongos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Fitoterapia , Extratos Vegetais/farmacologia , Caules de Planta
8.
Macromol Biosci ; 18(12): e1800335, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30408349

RESUMO

Surface topography has a profound effect on the development of the nervous system, such as neuronal differentiation and morphogenesis. While the interaction of neurons and the surface topography of their local environment is well characterized, the neuron-topography interaction during the regeneration process remains largely unknown. To address this question, an anisotropic surface topography resembling linear grooves made from poly(ethylene-vinyl acetate) (EVA), a soft and biocompatible polymer, using nanoimprinting, is established. It is found that neurons from both the central and peripheral nervous system can survive and grow on this grooved surface. Additionally, it is observed that axons but not dendrites specifically align with these grooves. Furthermore, it is demonstrated that neurons on the grooved surface are capable of regeneration after an on-site injury. More importantly, these injured neurons have an accelerated and enhanced regeneration. Together, the data demonstrate that this anisotropic topography guides axon growth and improves axon regeneration. This opens up the possibility to study the effect of surface topography on regenerating axons and has the potential to be developed into a medical device for treating peripheral nerve injuries.


Assuntos
Axônios/efeitos dos fármacos , Materiais Biocompatíveis/farmacologia , Regeneração Tecidual Guiada/métodos , Regeneração Nervosa/efeitos dos fármacos , Traumatismos dos Nervos Periféricos/terapia , Polietilenos/farmacologia , Polivinil/farmacologia , Animais , Anisotropia , Axônios/ultraestrutura , Materiais Biocompatíveis/síntese química , Materiais Biocompatíveis/química , Córtex Cerebral/citologia , Córtex Cerebral/efeitos dos fármacos , Gânglios Espinais/lesões , Gânglios Espinais/cirurgia , Camundongos , Camundongos Endogâmicos C57BL , Impressão Molecular/métodos , Nanoestruturas/química , Nanoestruturas/ultraestrutura , Regeneração Nervosa/fisiologia , Neuritos/efeitos dos fármacos , Neuritos/ultraestrutura , Traumatismos dos Nervos Periféricos/patologia , Polietilenos/síntese química , Polietilenos/química , Polivinil/síntese química , Polivinil/química , Cultura Primária de Células , Ratos , Células Receptoras Sensoriais/efeitos dos fármacos , Células Receptoras Sensoriais/ultraestrutura
9.
Sci Rep ; 8(1): 13197, 2018 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-30181617

RESUMO

Molecules and pathways that suppress growth are expressed in postmitotic neurons, a potential advantage in mature neural networks, but a liability during regeneration. In this work, we probed the APC (adenomatous polyposis coli)-ß-catenin partner pathway in adult peripheral sensory neurons during regeneration. APC had robust expression in the cytoplasm and perinuclear region of adult DRG sensory neurons both before and after axotomy injury. ß-catenin was expressed in neuronal nuclei, neuronal cytoplasm and also in perineuronal satellite cells. In injured dorsal root ganglia (DRG) sensory neurons and their axons, we observed paradoxical APC upregulation, despite its role as an inhibitor of growth whereas ß-catenin was downregulated. Inhibition of APC in adult sensory neurons and activation of ß-catenin, LEF/TCF transcriptional factors were associated with increased neuronal plasticity in vitro. Local knockdown of APC, at the site of sciatic nerve crush injury enhanced evidence for electrophysiological, behavioural and structural regeneration in vivo. This was accompanied by upregulation of ß-catenin. Collectively, the APC-ß-catenin-LEF/TCF transcriptional pathway impacts intrinsic mechanisms of axonal regeneration and neuronal plasticity after injury, offering new options for addressing axon regeneration.


Assuntos
Polipose Adenomatosa do Colo/metabolismo , Gânglios Espinais/fisiologia , Regeneração Nervosa , Transdução de Sinais , beta Catenina/metabolismo , Animais , Axônios , Células Cultivadas , Gânglios Espinais/citologia , Gânglios Espinais/lesões , Gânglios Espinais/patologia , Masculino , Ratos Sprague-Dawley , Células Receptoras Sensoriais/citologia , Células Receptoras Sensoriais/metabolismo , Células Receptoras Sensoriais/patologia
10.
Cell Rep ; 24(7): 1865-1879.e9, 2018 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-30110642

RESUMO

We generated a knockout mouse for the neuronal-specific ß-tubulin isoform Tubb3 to investigate its role in nervous system formation and maintenance. Tubb3-/- mice have no detectable neurobehavioral or neuropathological deficits, and upregulation of mRNA and protein of the remaining ß-tubulin isotypes results in equivalent total ß-tubulin levels in Tubb3-/- and wild-type mice. Despite similar levels of total ß-tubulin, adult dorsal root ganglia lacking TUBB3 have decreased growth cone microtubule dynamics and a decreased neurite outgrowth rate of 22% in vitro and in vivo. The effect of the 22% slower growth rate is exacerbated for sensory recovery, where fibers must reinnervate the full volume of the skin to recover touch function. Overall, these data reveal that, while TUBB3 is not required for formation of the nervous system, it has a specific role in the rate of peripheral axon regeneration that cannot be replaced by other ß-tubulins.


Assuntos
Regeneração Nervosa/genética , Crescimento Neuronal/genética , Isoformas de Proteínas/genética , Tubulina (Proteína)/genética , Potenciais de Ação/fisiologia , Animais , Feminino , Gânglios Espinais/lesões , Gânglios Espinais/metabolismo , Regulação da Expressão Gênica , Masculino , Aprendizagem em Labirinto , Camundongos , Camundongos Knockout , Microtúbulos/metabolismo , Microtúbulos/ultraestrutura , Plasticidade Neuronal/genética , Isoformas de Proteínas/metabolismo , Transdução de Sinais , Tubulina (Proteína)/deficiência
11.
Neurol Res ; 40(11): 930-937, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30088803

RESUMO

OBJECTIVE: Peripheral nerve injury (PNI) is a significant health problem that is linked to sensory, motor, and autonomic deficits. This pathological condition leads to a reduced quality of life in most affected individuals. Schwann cells (SCs) play a crucial role in the repair of PNI. Effective agents that promote SC activation may facilitate and accelerate peripheral nerve repair. Thymoquinone (TQ), a bioactive component of Nigella sativa seeds, has an antioxidant, anti-inflammatory, immunomodulatory, and neuroprotective properties. In the present study, the neuroprotective efficacy of TQ was investigated by using a laser microdissection technique in a mouse PNI model. METHODS: Single cells were isolated from dorsal root ganglions (DRGs) of 6-8-week-old mice, maintained in defined culture conditions and treated with or without TQ at different concentrations. Axons were cut (axotomy) using a controllable laser microbeam to model axonal injury in vitro. Under fluorescence microscopy, cell viability was evaluated using the fluorescent dyes. The behavior of the cells was continuously monitored with time-lapse video microscopy. RESULTS: TQ significantly increased neuronal survival by promoting the survival and proliferation of SCs and fibroblasts, as well as the migration of SCs. Furthermore, TQ improved the ability to extend neurites of axotomized neurons. The regenerative effect of TQ was dose-dependent suggesting a target specificity. Our studies warrant further preclinical and clinical investigations of TQ as a potential regenerative agent to treat peripheral nerve injuries. CONCLUSION: TQ exhibits a regenerative potential for the treatment of damaged peripheral nerves.


Assuntos
Benzoquinonas/farmacologia , Morte Celular/efeitos dos fármacos , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/lesões , Fármacos Neuroprotetores/farmacologia , Animais , Axônios/efeitos dos fármacos , Axônios/patologia , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Fibroblastos/efeitos dos fármacos , Fibroblastos/patologia , Gânglios Espinais/patologia , Masculino , Camundongos Endogâmicos BALB C , Regeneração Nervosa/efeitos dos fármacos , Crescimento Neuronal/efeitos dos fármacos , Traumatismos dos Nervos Periféricos/tratamento farmacológico , Traumatismos dos Nervos Periféricos/patologia , Células de Schwann/efeitos dos fármacos , Células de Schwann/patologia
12.
Braz J Med Biol Res ; 51(10): e7113, 2018 Jul 26.
Artigo em Inglês | MEDLINE | ID: mdl-30066726

RESUMO

Dorsal root ganglia (DRG) neurons regenerate spontaneously after traumatic or surgical injury. Long noncoding RNAs (lncRNAs) are involved in various biological regulation processes. Conditions of lncRNAs in DRG neuron injury deserve to be further investigated. Transcriptomic analysis was performed by high-throughput Illumina HiSeq2500 sequencing to profile the differential genes in L4-L6 DRGs following rat sciatic nerve tying. A total of 1,228 genes were up-regulated and 1,415 down-regulated. By comparing to rat lncRNA database, 86 known and 26 novel lncRNA genes were found to be differential. The 86 known lncRNA genes modulated 866 target genes subject to gene ontology (GO) and KEGG enrichment analysis. The genes involved in the neurotransmitter status of neurons were downregulated and those involved in a neuronal regeneration were upregulated. Known lncRNA gene rno-Cntnap2 was downregulated. There were 13 credible GO terms for the rno-Cntnap2 gene, which had a putative function in cell component of voltage-gated potassium channel complex on the cell surface for neurites. In 26 novel lncRNA genes, 4 were related to 21 mRNA genes. A novel lncRNA gene AC111653.1 improved rno-Hypm synthesizing huntingtin during sciatic nerve regeneration. Real time qPCR results attested the down-regulation of rno-Cntnap lncRNA gene and the upregulation of AC111653.1 lncRNA gene. A total of 26 novel lncRNAs were found. Known lncRNA gene rno-Cntnap2 and novel lncRNA AC111653.1 were involved in neuropathic pain of DRGs after spared sciatic nerve injury. They contributed to peripheral nerve regeneration via the putative mechanisms.


Assuntos
Gânglios Espinais/lesões , Neuralgia/metabolismo , Traumatismos dos Nervos Periféricos/metabolismo , RNA Longo não Codificante/metabolismo , RNA Mensageiro/genética , Nervo Isquiático/metabolismo , Transcriptoma , Animais , Sequência de Bases , Western Blotting , Mapeamento Cromossômico , Modelos Animais de Doenças , Gânglios Espinais/metabolismo , Gânglios Espinais/fisiopatologia , Regulação da Expressão Gênica , Masculino , Dados de Sequência Molecular , Neuralgia/genética , Neuralgia/fisiopatologia , Traumatismos dos Nervos Periféricos/genética , Traumatismos dos Nervos Periféricos/fisiopatologia , RNA Longo não Codificante/genética , RNA Longo não Codificante/fisiologia , RNA Mensageiro/metabolismo , RNA Mensageiro/fisiologia , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Nervo Isquiático/fisiopatologia
13.
Curr Protoc Stem Cell Biol ; 47(1): e58, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30021049

RESUMO

Human mesenchymal stromal stem cells (hMSCs) hold regenerative medicine potential due to their availability, in vitro expansion readiness, and autologous feasibility. For neural repair, hMSCs show translational value in research on stroke, spinal cord injury (SCI), and traumatic brain injury. It is pivotal to establish multimodal in vitro systems to investigate molecular mechanisms underlying neural actions of hMSCs. Here, we describe a platform protocol on how to set up organotypic co-cultures of hMSCs (alone or polymer-scaffolded) with explanted adult rat dorsal root ganglia (DRGs) to determine neural injury and recovery events for designing implants to counteract neurotrauma sequelae. We emphasize in vitro hMSC propagation, polymer scaffolding, hMSC stemness maintenance, hMSC-DRG interaction profiling, and analytical formulas of neuroinflammation, trophic factor expression, DRG neurite outgrowth and tropic tracking, and in vivo verification of tailored implants in rodent models of SCI. © 2018 by John Wiley & Sons, Inc.


Assuntos
Técnicas de Cocultura/métodos , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Traumatismos do Sistema Nervoso/terapia , Animais , Gânglios Espinais/citologia , Gânglios Espinais/lesões , Humanos , Ratos
14.
J Recept Signal Transduct Res ; 38(3): 198-203, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29932348

RESUMO

OBJECTIVES: Here, using rat model, we investigated the roles of gardenoside in the chronic constriction injury (CCI) of the ischiadic nerve. METHODS: Bennett and Xie's unilateral sciatic nerve CCI model was used in this study. A total of 60 rats were divided into control group (CN), sham group (Sham), CCI group, and gardenoside administrated CCI group. An aliquot of 5 mL gardenoside solution was administrated through gavage once per day for 14 d. Mechanical withdrawal threshold (MWT) and the thermal withdrawal latency (TWL) were detected. The levels of inducible nitric oxide synthase (iNOS), interleukin-1ß (IL-1ß), and tumor necrosis factor-α (TNF-α) in spinal fluid were detected by ELISA. By using real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blot, we analyzed the expression of P2X purinoceptor 3 and 7 (P2X3 and P2X7 receptors) in different groups. The expression of p-ERK/ERK and p-p38/p38 were also detected by western blot. RESULTS: We found out that gardenoside could significantly improve the sciatica by partially restore the decrease of MWT and TWL in CCI rats. The levels of iNOS, IL-1ß, and TNF-α were higher in CCI group (p < .05). The expressions of P2X3 and P2X7 were significantly increased in the CCI rats compared to control rats (p < .05). The levels of p-ERK/ERK and p-p38/p38 were also obviously increased in CCI group (p < .05). After treated with the gardenoside, these increases were decreased. CONCLUSIONS: These results indicated that gardenoside may be able to relief CCI-induced neuropathic pain by regulating the P2X3 and the P2X7 expression on the ischiadic nerve.


Assuntos
Iridoides/administração & dosagem , Neuralgia/tratamento farmacológico , Receptores Purinérgicos P2X3/genética , Receptores Purinérgicos P2X7/genética , Animais , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/lesões , Gânglios Espinais/patologia , Regulação da Expressão Gênica , Humanos , Interleucina-1beta , Neuralgia/genética , Neuralgia/fisiopatologia , Óxido Nítrico Sintase Tipo II , Limiar da Dor , Ratos , Nervo Isquiático/efeitos dos fármacos , Nervo Isquiático/lesões , Nervo Isquiático/fisiopatologia , Fator de Necrose Tumoral alfa
15.
J Neurosci ; 38(27): 6090-6101, 2018 07 04.
Artigo em Inglês | MEDLINE | ID: mdl-29875269

RESUMO

Nerve injury-induced hyperactivity of primary sensory neurons in the dorsal root ganglion (DRG) contributes to chronic pain development, but the underlying epigenetic mechanisms remain poorly understood. Here we determined genome-wide changes in DNA methylation in the nervous system in neuropathic pain. Spinal nerve ligation (SNL), but not paclitaxel treatment, in male Sprague Dawley rats induced a consistent low-level hypomethylation in the CpG sites in the DRG during the acute and chronic phases of neuropathic pain. DNA methylation remodeling in the DRG occurred early after SNL and persisted for at least 3 weeks. SNL caused DNA methylation changes at 8% of CpG sites with prevailing hypomethylation outside of CpG islands, in introns, intergenic regions, and repetitive sequences. In contrast, SNL caused more gains of methylation in the spinal cord and prefrontal cortex. The DNA methylation changes in the injured DRGs recapitulated developmental reprogramming at the neonatal stage. Methylation reprogramming was correlated with increased gene expression variability. A diet deficient in methyl donors induced hypomethylation and pain hypersensitivity. Intrathecal administration of the DNA methyltransferase inhibitor RG108 caused long-lasting pain hypersensitivity. DNA methylation reprogramming in the DRG thus contributes to nerve injury-induced chronic pain. Restoring DNA methylation may represent a new therapeutic approach to treat neuropathic pain.SIGNIFICANCE STATEMENT Epigenetic mechanisms are critically involved in the transition from acute to chronic pain after nerve injury. However, genome-wide changes in DNA methylation in the nervous system and their roles in neuropathic pain development remain unclear. Here we used digital restriction enzyme analysis of methylation to quantitatively determine genome-wide DNA methylation changes caused by nerve injury. We showed that nerve injury caused DNA methylation changes at 8% of CpG sites with prevailing hypomethylation outside of CpG islands in the dorsal root ganglion. Reducing DNA methylation induced pain hypersensitivity, whereas increasing DNA methylation attenuated neuropathic pain. These findings extend our understanding of the epigenetic mechanism of chronic neuropathic pain and suggest new strategies to treat nerve injury-induced chronic pain.


Assuntos
Dor Crônica/metabolismo , Metilação de DNA/fisiologia , Gânglios Espinais/metabolismo , Neuralgia/metabolismo , Animais , Dor Crônica/genética , Epigênese Genética/genética , Gânglios Espinais/lesões , Masculino , Neuralgia/genética , Ratos , Ratos Sprague-Dawley
16.
Science ; 359(6382): 1416-1421, 2018 03 23.
Artigo em Inglês | MEDLINE | ID: mdl-29567716

RESUMO

How is protein synthesis initiated locally in neurons? We found that mTOR (mechanistic target of rapamycin) was activated and then up-regulated in injured axons, owing to local translation of mTOR messenger RNA (mRNA). This mRNA was transported into axons by the cell size-regulating RNA-binding protein nucleolin. Furthermore, mTOR controlled local translation in injured axons. This included regulation of its own translation and that of retrograde injury signaling molecules such as importin ß1 and STAT3 (signal transducer and activator of transcription 3). Deletion of the mTOR 3' untranslated region (3'UTR) in mice reduced mTOR in axons and decreased local translation after nerve injury. Both pharmacological inhibition of mTOR in axons and deletion of the mTOR 3'UTR decreased proprioceptive neuronal survival after nerve injury. Thus, mRNA localization enables spatiotemporal control of mTOR pathways regulating local translation and long-range intracellular signaling.


Assuntos
Axônios/metabolismo , Gânglios Espinais/lesões , Biossíntese de Proteínas , Nervo Isquiático/lesões , Serina-Treonina Quinases TOR/biossíntese , Regiões 3' não Traduzidas , Animais , Tamanho Celular , Camundongos , Camundongos Endogâmicos , Fosfoproteínas/metabolismo , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Ratos , Ratos Endogâmicos BB , Ratos Sprague-Dawley , Transdução de Sinais , Serina-Treonina Quinases TOR/genética
17.
Nat Cell Biol ; 20(3): 307-319, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29434374

RESUMO

Reactive oxygen species (ROS) contribute to tissue damage and remodelling mediated by the inflammatory response after injury. Here we show that ROS, which promote axonal dieback and degeneration after injury, are also required for axonal regeneration and functional recovery after spinal injury. We find that ROS production in the injured sciatic nerve and dorsal root ganglia requires CX3CR1-dependent recruitment of inflammatory cells. Next, exosomes containing functional NADPH oxidase 2 complexes are released from macrophages and incorporated into injured axons via endocytosis. Once in axonal endosomes, active NOX2 is retrogradely transported to the cell body through an importin-ß1-dynein-dependent mechanism. Endosomal NOX2 oxidizes PTEN, which leads to its inactivation, thus stimulating PI3K-phosporylated (p-)Akt signalling and regenerative outgrowth. Challenging the view that ROS are exclusively involved in nerve degeneration, we propose a previously unrecognized role of ROS in mammalian axonal regeneration through a NOX2-PI3K-p-Akt signalling pathway.


Assuntos
Axônios/enzimologia , Exossomos/enzimologia , Gânglios Espinais/enzimologia , NADPH Oxidase 2/metabolismo , Degeneração Neural , Regeneração Nervosa , Traumatismos dos Nervos Periféricos/enzimologia , Espécies Reativas de Oxigênio/metabolismo , Nervo Isquiático/enzimologia , Traumatismos da Medula Espinal/enzimologia , Animais , Axônios/patologia , Receptor 1 de Quimiocina CX3C/metabolismo , Linhagem Celular , Modelos Animais de Doenças , Dineínas/metabolismo , Endocitose , Endossomos/enzimologia , Endossomos/patologia , Exossomos/patologia , Gânglios Espinais/lesões , Gânglios Espinais/patologia , Macrófagos/enzimologia , Macrófagos/patologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , NADPH Oxidase 2/deficiência , NADPH Oxidase 2/genética , Proteínas Nucleares/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Traumatismos dos Nervos Periféricos/genética , Traumatismos dos Nervos Periféricos/patologia , Traumatismos dos Nervos Periféricos/fisiopatologia , Fosfatidilinositol 3-Quinase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Nervo Isquiático/lesões , Nervo Isquiático/patologia , Nervo Isquiático/fisiopatologia , Transdução de Sinais , Traumatismos da Medula Espinal/genética , Traumatismos da Medula Espinal/patologia , Traumatismos da Medula Espinal/fisiopatologia , beta Carioferinas
18.
World Neurosurg ; 114: e35-e41, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29408594

RESUMO

OBJECTIVE: To explore the effect of the inhibitor of histone deacetylase (6HDAC6), tubastatin A, on the functional recovery of injured central branch of dorsal root ganglia (cauda equina). METHODS: A total of 30 Sprague-Dawley rats (n = 10 for each group) were divided randomly into sham operation (sham group), cauda equina compression control (CEC control group), and cauda equina compression plus tubastatin A treatment (tubastatin A group). The tail-flick test was performed to detect the sense of pain and warmth as well as motor function. Immunoblotting/immunofluorescence experiments, terminal deoxynucleotidyl transferase dUTP nick end labeling staining, and hematoxylin-eosin staining were performed to detect the amount of HDAC6 in dorsal root ganglion (DRG) neurons, degree of apoptosis in DRG neurons, and degree of cauda equina injury, respectively. RESULTS: The ratio of apoptotic cells in the CEC control group was greater than that in the sham group, whereas it decreased in the tubastatin A group. Hematoxylin-eosin staining revealed that the fibers of cauda equina in the tubastatin A group were more compact compared with those in the CEC control group. The expression of HDAC6 was not different between the sham and CEC control groups, whereas it decreased significantly in the tubastatin A group. Tubastatin A administration shortened tail-flick latency on the seventh day after operation compared with the CEC control group. CONCLUSIONS: Tubastatin A significantly decreased the expression of HDAC6 in DRG neurons with injured cauda equina, inhibited the apoptosis of neural cells and axonal demyelinating changes in cauda equina, and partially promoted the recovery of neural function.


Assuntos
Gânglios Espinais/efeitos dos fármacos , Ácidos Hidroxâmicos/farmacologia , Indóis/farmacologia , Recuperação de Função Fisiológica/efeitos dos fármacos , Estenose Espinal/tratamento farmacológico , Animais , Gânglios Espinais/lesões , Gânglios Espinais/fisiopatologia , Desacetilase 6 de Histona/metabolismo , Vértebras Lombares/efeitos dos fármacos , Vértebras Lombares/fisiopatologia , Masculino , Neurônios/efeitos dos fármacos , Traumatismos dos Nervos Periféricos/tratamento farmacológico , Traumatismos dos Nervos Periféricos/fisiopatologia , Radiculopatia/tratamento farmacológico , Ratos Sprague-Dawley , Estenose Espinal/fisiopatologia
19.
Cancer Res ; 78(3): 817-829, 2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29191802

RESUMO

Chemotherapy-induced peripheral neuropathy (CIPN) is a major cause of disability in cancer survivors. CIPN investigations in preclinical model systems have focused on either behaviors or acute changes in nerve conduction velocity (NCV) and amplitude, but greater understanding of the underlying nature of axonal injury and its long-term processes is needed as cancer patients live longer. In this study, we used multiple independent endpoints to systematically characterize CIPN recovery in mice exposed to the antitubulin cancer drugs eribulin, ixabepilone, paclitaxel, or vinorelbine at MTDs. All of the drugs ablated intraepidermal nerve fibers and produced axonopathy, with a secondary disruption in myelin structure within 2 weeks of drug administration. In addition, all of the drugs reduced sensory NCV and amplitude, with greater deficits after paclitaxel and lesser deficits after ixabepilone. These effects correlated with degeneration in dorsal root ganglia (DRG) and sciatic nerve and abundance of Schwann cells. Although most injuries were fully reversible after 3-6 months after administration of eribulin, vinorelbine, and ixabepilone, we observed delayed recovery after paclitaxel that produced a more severe, pervasive, and prolonged neurotoxicity. Compared with other agents, paclitaxel also displayed a unique prolonged exposure in sciatic nerve and DRG. The most sensitive indicator of toxicity was axonopathy and secondary myelin changes accompanied by a reduction in intraepidermal nerve fiber density. Taken together, our findings suggest that intraepidermal nerve fiber density and changes in NCV and amplitude might provide measures of axonal injury to guide clinical practice.Significance: This detailed preclinical study of the long-term effects of widely used antitubulin cancer drugs on the peripheral nervous system may help guide clinical evaluations to improve personalized care in limiting neurotoxicity in cancer survivors. Cancer Res; 78(3); 817-29. ©2017 AACR.


Assuntos
Gânglios Espinais/efeitos dos fármacos , Microtúbulos/efeitos dos fármacos , Doenças do Sistema Nervoso Periférico/induzido quimicamente , Recuperação de Função Fisiológica/efeitos dos fármacos , Células de Schwann/efeitos dos fármacos , Nervo Isquiático/efeitos dos fármacos , Moduladores de Tubulina/toxicidade , Doença Aguda , Animais , Células Cultivadas , Feminino , Gânglios Espinais/lesões , Gânglios Espinais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Microtúbulos/patologia , Doenças do Sistema Nervoso Periférico/patologia , Células de Schwann/patologia , Nervo Isquiático/lesões , Nervo Isquiático/patologia
20.
Colloids Surf B Biointerfaces ; 162: 126-134, 2018 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-29190463

RESUMO

Schwann cell-seeded nerve guidance channels are designed to assist post-traumatic nerve regeneration in the PNS. Chitosan is a natural polymer well suited for tissue engineering as it is biocompatible, non-immunogenic, and biodegradable. Electrospun chitosan nanofibers utilized in nerve guidance channels have the capacity for guiding axonal growth within the channel lumen yet are limited in their capacity to maintain structural integrity within physiological environments. To address this, we attempted genipin crosslinking of chitosan nanofibers. Compared to neat chitosan nanofibers, genipin-treated nanofibers exhibited increased stiffness, resistance to swelling and lysozymal degradation. Furthermore, alignment and proliferation of purified Schwann cell cultures upon genipin-treated substratum was enhanced. When dorsal root ganglion explants were utilized as an in vitro model of peripheral nerve regeneration, emigrating neurons and Schwann cells assumed the uniaxial pattern of aligned electrospun chitosan nanofibers. Neurite growth along the nanofibers led, reaching a frontier more than twice that of the pursuant Schwann cells. Critically, neurite growth rate upon genipin-treated nanofibers demonstrated a 100% increase. Altogether, genipin treatment improves upon the physical and biological properties of chitosan nanofibers towards their utility in nerve guidance channel design.


Assuntos
Quitosana/farmacologia , Iridoides/química , Regeneração Nervosa/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Células de Schwann/efeitos dos fármacos , Engenharia Tecidual/métodos , Animais , Materiais Biocompatíveis/química , Materiais Biocompatíveis/farmacologia , Quitosana/química , Reagentes para Ligações Cruzadas/química , Gânglios Espinais/citologia , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/lesões , Nanofibras/química , Nanofibras/ultraestrutura , Crescimento Neuronal/efeitos dos fármacos , Neurônios/citologia , Cultura Primária de Células , Ratos , Ratos Sprague-Dawley , Células de Schwann/citologia , Nervo Isquiático/citologia , Nervo Isquiático/efeitos dos fármacos , Nervo Isquiático/lesões , Técnicas de Cultura de Tecidos , Tecidos Suporte
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