RESUMEN
Synaptic changes are early manifestations of neuronal dysfunction in Huntington's disease (HD). However, the mechanisms by which mutant HTT protein impacts synaptogenesis and function are not well understood. Herein we explored HD pathogenesis in the BACHD mouse model by examining synaptogenesis and function in long term primary cortical cultures. At DIV14 (days in vitro), BACHD cortical neurons showed no difference from WT neurons in synaptogenesis as revealed by colocalization of a pre-synaptic (Synapsin I) and a post-synaptic (PSD95) marker. From DIV21 to DIV35, BACHD neurons showed progressively reduced colocalization of Synapsin I and PSD95 relative to WT neurons. The deficits were effectively rescued by treatment of BACHD neurons with BDNF. The recombinant apical domain of CCT1 (ApiCCT1) yielded a partial rescuing effect. BACHD neurons also showed culture age-related significant functional deficits as revealed by multielectrode arrays (MEAs). These deficits were prevented by BDNF, whereas ApiCCT1 showed a less potent effect. These findings are evidence that deficits in BACHD synapse and function can be replicated in vitro and that BDNF or a TRiC-inspired reagent can potentially be protective against these changes in BACHD neurons. Our findings support the use of cellular models to further explicate HD pathogenesis and potential treatments.
Asunto(s)
Factor Neurotrófico Derivado del Encéfalo , Corteza Cerebral , Modelos Animales de Enfermedad , Enfermedad de Huntington , Neuronas , Sinapsis , Animales , Enfermedad de Huntington/metabolismo , Enfermedad de Huntington/patología , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Sinapsis/metabolismo , Sinapsis/efectos de los fármacos , Sinapsis/patología , Corteza Cerebral/metabolismo , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/patología , Ratones , Neuronas/metabolismo , Neuronas/efectos de los fármacos , Neuronas/patología , Ratones Transgénicos , Células Cultivadas , Sinapsinas/metabolismo , Proteína Huntingtina/genética , Proteína Huntingtina/metabolismo , Ratones Endogámicos C57BLRESUMEN
Huntington's disease (HD) is caused by an expanded CAG repeat in the huntingtin gene, yielding a Huntingtin protein with an expanded polyglutamine tract. While experiments with patient-derived induced pluripotent stem cells (iPSCs) can help understand disease, defining pathological biomarkers remains challenging. Here, we used cryogenic electron tomography to visualize neurites in HD patient iPSC-derived neurons with varying CAG repeats, and primary cortical neurons from BACHD, deltaN17-BACHD, and wild-type mice. In HD models, we discovered sheet aggregates in double membrane-bound organelles, and mitochondria with distorted cristae and enlarged granules, likely mitochondrial RNA granules. We used artificial intelligence to quantify mitochondrial granules, and proteomics experiments reveal differential protein content in isolated HD mitochondria. Knockdown of Protein Inhibitor of Activated STAT1 ameliorated aberrant phenotypes in iPSC- and BACHD neurons. We show that integrated ultrastructural and proteomic approaches may uncover early HD phenotypes to accelerate diagnostics and the development of targeted therapeutics for HD.
Asunto(s)
Enfermedad de Huntington , Células Madre Pluripotentes Inducidas , Animales , Ratones , Inteligencia Artificial , Modelos Animales de Enfermedad , Proteína Huntingtina/genética , Proteína Huntingtina/metabolismo , Enfermedad de Huntington/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Mitocondrias/metabolismo , Neuronas/metabolismo , Fenotipo , Proteómica , HumanosRESUMEN
Mutations or triplication of the alpha synuclein (ASYN) gene contribute to synucleinopathies including Parkinson's disease (PD), Dementia with Lewy bodies (DLB) and multiple system atrophy (MSA). Recent evidence suggests that ASYN also plays an important role in amyloid-induced neurotoxicity, although the mechanism(s) remains unknown. One hypothesis is that accumulation of ASYN alters endolysosomal pathways to impact axonal trafficking and processing of the amyloid precursor protein (APP). To define an axonal function for ASYN, we used a transgenic mouse model of synucleinopathy that expresses a GFP-human ASYN (GFP-hASYN) transgene and an ASYN knockout (ASYN-/-) mouse model. Our results demonstrate that expression of GFP-hASYN in primary neurons derived from a transgenic mouse impaired axonal trafficking and processing of APP. In addition, axonal transport of BACE1, Rab5, Rab7, lysosomes and mitochondria were also reduced in these neurons. Interestingly, axonal transport of these organelles was also affected in ASYN-/- neurons, suggesting that ASYN plays an important role in maintaining normal axonal transport function. Therefore, selective impairment of trafficking and processing of APP by ASYN may act as a potential mechanism to induce pathological features of Alzheimer's disease (AD) in PD patients.
Asunto(s)
Enfermedad de Parkinson , Sinucleinopatías , Humanos , Ratones , Animales , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismo , Secretasas de la Proteína Precursora del Amiloide , Precursor de Proteína beta-Amiloide/genética , Ácido Aspártico Endopeptidasas , Enfermedad de Parkinson/genética , Ratones Transgénicos , Lisosomas/metabolismoRESUMEN
Rab7 GTPase regulates mitochondrial morphology and function. Missense mutation(s) of Rab7 underlies the pathogenesis of Charcot Marie Tooth 2B (CMT2B) peripheral neuropathy. Herein, we investigate how mitochondrial morphology and function are impacted by the CMT2B associated Rab7V162M mutation. In contrast to recent studies of using heterologous overexpression systems, our results demonstrate significant mitochondrial fragmentation in both human CMT2B patient fibroblasts and CMT2B embryonic fibroblasts (MEFs). Primary cultured E18 dorsal root ganglion (DRG) sensory neurons also show mitochondrial fragmentation and altered axonal mitochondrial movement. In addition, we demonstrate that inhibitors to either the mitochondrial fission protein Drp1 or to the nucleotide binding to Rab7 normalize the mitochondrial deficits in both MEFs and E18 cultured DRG neurons. Our study reveals, for the first time, that expression of CMT2B Rab7 mutation at the physiological level enhances Drp1 activity to promote mitochondrial fission, potentially underlying selective vulnerability of peripheral sensory neurons in CMT2B pathogenesis.
Asunto(s)
Enfermedad de Charcot-Marie-Tooth , Proteínas de Unión al GTP rab , Enfermedad de Charcot-Marie-Tooth/genética , Humanos , Laminopatías , Mitocondrias/metabolismo , Células Receptoras Sensoriales/metabolismo , Proteínas de Unión al GTP rab/genética , Proteínas de Unión al GTP rab/metabolismo , Proteínas de Unión a GTP rab7RESUMEN
Laser-induced shockwaves (LIS) can be utilized as a method to subject cells to conditions similar to those occurring during a blast-induced traumatic brain injury. The pairing of LIS with genetically encoded biosensors allows researchers to monitor the immediate molecular events resulting from such an injury. In this study, we utilized the genetically encoded Ca2+ FRET biosensor D3CPV to study the immediate Ca2+ response to laser-induced shockwave in cortical neurons and Schwann cells. Our results show that both cell types exhibit a transient Ca2+ increase irrespective of extracellular Ca2+ conditions. LIS allows for the simultaneous monitoring of the effects of shear stress on cells, as well as nearby cell damage and death.
RESUMEN
Alzheimer's disease (AD) is associated with the accumulation and deposition of a beta-amyloid (Αß) peptide in the brain, resulting in increased neuroinflammation and synaptic dysfunction. Intranasal delivery of targeted drugs to the brain represents a noninvasive pathway that bypasses the blood-brain barrier and minimizes systemic exposure. The aim of this study was to evaluate the therapeutic effect of intranasally delivered 9-cis retinoic acid (RA) on the neuropathology of an AD mouse model. Herein, we observed dramatically decreased Αß deposition in the brains of amyloid precursor protein (APP) and presenilin 1 (PS1) double-transgenic mice (APP/PS1) treated intranasally with 9-cis RA for 4 weeks compared to that in the brains of vehicle-treated mice. Importantly, intranasal delivery of 9-cis RA suppressed Αß-associated astrocyte activation and neuroinflammation and ultimately restored synaptic deficits in APP/PS1 transgenic mice. These results support the critical roles of Αß-associated neuroinflammation responses to synaptic deficits, particularly during the deposition of Αß. Our findings provide strong evidence that intranasally delivered 9-cis RA attenuates neuronal dysfunction in an AD mouse model and is a promising therapeutic strategy for the prevention and treatment of AD.
Asunto(s)
Alitretinoína/farmacología , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/metabolismo , Mediadores de Inflamación/metabolismo , Inflamación/metabolismo , Administración Intranasal , Alitretinoína/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Animales , Astrocitos/metabolismo , Astrocitos/patología , Encéfalo/metabolismo , Modelos Animales de Enfermedad , Ratones , Ratones Transgénicos , Microglía/patología , Presenilina-1RESUMEN
Nerve growth factor (NGF) exerts multiple functions on target neurons throughout development. The recent discovery of a point mutation leading to a change from arginine to tryptophan at residue 100 in the mature NGFß sequence (NGFR100W) in patients with hereditary sensory and autonomic neuropathy type V (HSAN V) made it possible to distinguish the signaling mechanisms that lead to two functionally different outcomes of NGF: trophic versus nociceptive. We performed extensive biochemical, cellular, and live-imaging experiments to examine the binding and signaling properties of NGFR100W Our results show that, similar to the wild-type NGF (wtNGF), the naturally occurring NGFR100W mutant was capable of binding to and activating the TrkA receptor and its downstream signaling pathways to support neuronal survival and differentiation. However, NGFR100W failed to bind and stimulate the 75 kDa neurotrophic factor receptor (p75NTR)-mediated signaling cascades (i.e., the RhoA-Cofilin pathway). Intraplantar injection of NGFR100W into adult rats induced neither TrkA-mediated thermal nor mechanical acute hyperalgesia, but retained the ability to induce chronic hyperalgesia based on agonism for TrkA signaling. Together, our studies provide evidence that NGFR100W retains trophic support capability through TrkA and one aspect of its nociceptive signaling, but fails to engage p75NTR signaling pathways. Our findings suggest that wtNGF acts via TrkA to regulate the delayed priming of nociceptive responses. The integration of both TrkA and p75NTR signaling thus appears to regulate neuroplastic effects of NGF in peripheral nociception.SIGNIFICANCE STATEMENT In the present study, we characterized the naturally occurring nerve growth factor NGFR100W mutant that is associated with hereditary sensory and autonomic neuropathy type V. We have demonstrated for the first time that NGFR100W retains trophic support capability through TrkA, but fails to engage p75NTR signaling pathways. Furthermore, after intraplantar injection into adult rats, NGFR100W induced neither thermal nor mechanical acute hyperalgesia, but retained the ability to induce chronic hyperalgesia. We have also provided evidence that the integration of both TrkA- and p75NTR-mediated signaling appears to regulate neuroplastic effects of NGF in peripheral nociception. Our study with NGFR100W suggests that it is possible to uncouple trophic effect from nociceptive function, both induced by wild-type NGF.
Asunto(s)
Neuropatías Hereditarias Sensoriales y Autónomas/genética , Mutación Missense , Factor de Crecimiento Nervioso/genética , Nocicepción , Receptor trkA/metabolismo , Receptores de Factor de Crecimiento Nervioso/metabolismo , Células 3T3 , Animales , Células Cultivadas , Células HEK293 , Neuropatías Hereditarias Sensoriales y Autónomas/metabolismo , Neuropatías Hereditarias Sensoriales y Autónomas/fisiopatología , Humanos , Masculino , Ratones , Factor de Crecimiento Nervioso/metabolismo , Proteínas del Tejido Nervioso , Células PC12 , Unión Proteica , Ratas , Ratas Sprague-Dawley , Receptores de Factores de Crecimiento , Transducción de SeñalRESUMEN
The objective of this study was to evaluate the efficacy and safety of intra-arterial thrombolysis in treating acute cerebral infarction and further discuss the indications of acute cerebral infarction treatment, in order to enhance the therapeutic effects of arterial thrombolysis. The data of 164 patients with acute cerebral infarction who accepted intra-arterial thrombolysis treatment by using rt-PA or reteplase between 2009 and 2014 at the Department of Neurology of our hospital, were collected, including patients' medical history, characteristics of the onset procedure, intervals between onset and intra-arterial thrombolysis, bleeding or death, and the changing process of patient's main neurologic function after the treatment. The neurological functions including muscle strength, speech, and level of consciousness were chosen for evaluation. Through a review of cerebral angiography, we collected the digital subtraction angiography (DSA) morphological changes of blood vessels before and after arterial thrombolysis to evaluate whether those blood vessels had been reperfused. Thereafter, we analyzed and statistically processed above-mentioned data. The mean time of arterial thrombolysis was 5.7 h. DSA results were as follows: 22 patients had complete internal carotid artery (ICA) occlusion; 49 patients middle cerebral artery's (MCA's) Ml or M2 segment occlusion; 6 patients anterior cerebral artery (ACA) occlusion; 58 patients reperfusion after thrombolysis, and the recanalization rate was 76 %. Based on vertebral-basilar artery (VBA) system, 18 patients had complete occlusion, 11 patients had reperfusion after thrombolysis, and the recanalization rate was 61 %. A total of 63 patients had severe stenosis, and they had significantly improved after thrombolysis. The clinical symptoms of patients were improved: 79 out of 164 patients with paralysis had partially recovered their limb muscle strength after operation, while 33 patients had completely recovered, and there was no recovery at all of the muscle strength in 4 patients after operation. In total, 59 out of 63 patients with aphasia had improved their language function, while 19 patients with disturbance of consciousness turned for the better after arterial thrombolysis. Only one patient experienced the cerebral hemorrhage, and 14 cases had gingival bleeding, oral mucosa bleeding, and urethrorrhagia. The overall effective rates of intra-arterial thrombolysis in treating the acute cerebral infarction by reteplase had no significant differences compared to those by rt-PA, and there were no hemorrhagic complications. It is safe and effective if the arterial thrombolysis using reteplase is performed within a few hours after acute cerebral infarction onset because reteplase has a higher clinical efficacy and lower hemorrhagic transformation, which suggests that it may become a new feasible option for clinical arterial thrombolysis.
Asunto(s)
Infarto Cerebral/tratamiento farmacológico , Fibrinolíticos/uso terapéutico , Adulto , Anciano , Anciano de 80 o más Años , Arterias Carótidas/diagnóstico por imagen , Angiografía Cerebral , Infarto Cerebral/diagnóstico por imagen , Femenino , Humanos , Infusiones Intraarteriales , Masculino , Persona de Mediana Edad , Arteria Cerebral Media/diagnóstico por imagen , Proteínas Recombinantes/uso terapéutico , Activador de Tejido Plasminógeno/genética , Activador de Tejido Plasminógeno/metabolismo , Activador de Tejido Plasminógeno/uso terapéuticoRESUMEN
BACKGROUND: Acid-base imbalance in various metabolic disturbances leads to human brain dysfunction. Compared with acidosis, the patients suffered from alkalosis demonstrate more severe neurological signs that are difficultly corrected. We hypothesize a causative process that the nerve cells in the brain are more vulnerable to alkalosis than acidosis. METHODS: The vulnerability of GABAergic neurons to alkalosis versus acidosis was compared by analyzing their functional changes in response to the extracellular high pH and low pH. The neuronal and synaptic functions were recorded by whole-cell recordings in the cortical slices. RESULTS: The elevation or attenuation of extracellular pH impaired these GABAergic neurons in terms of their capability to produce spikes, their responsiveness to excitatory synaptic inputs and their outputs via inhibitory synapses. Importantly, the dysfunction of these active properties appeared severer in alkalosis than acidosis. CONCLUSIONS: The severer impairment of cortical GABAergic neurons in alkalosis patients leads to more critical neural excitotoxicity, so that alkalosis-induced brain dysfunction is difficultly corrected, compared to acidosis. The vulnerability of cortical GABAergic neurons to high pH is likely a basis of severe clinical outcomes in alkalosis versus acidosis.
Asunto(s)
Acidosis/patología , Alcalosis/patología , Corteza Cerebral/patología , Neuronas GABAérgicas/patología , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/genética , Animales , Glutamato Descarboxilasa/genética , Glutamato Descarboxilasa/metabolismo , Proteínas Fluorescentes Verdes/genética , Técnicas In Vitro , Potenciales Postsinápticos Inhibidores/efectos de los fármacos , Potenciales Postsinápticos Inhibidores/genética , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Técnicas de Placa-ClampRESUMEN
The dopamine D3 receptor (DRD3) Ser9Gly variant has attracted more attention since the variant was observed to be associated with risk of essential tremor (ET). A number of association studies concerning the DRD3 Ser9Gly variant and ET susceptibility have been conducted in various populations. However, some results were contradictory. To derive a more precise estimation of the relationship between the DRD3 Ser9Gly variant and the genetic risk of ET, we performed a comprehensive meta-analysis which included seven case-control studies. The meta-analysis was conducted in four genetic models: dominant, recessive, heterozygous, and homozygous. The odds ratio and 95% confidence intervals were used as the measure of association. The combined results of overall analysis showed a lack of association of the DRD3 Ser9Gly variant and ET, regardless of the genetic model of Ser9Gly. Publication bias and heterogeneity were absent in most analyses. In conclusion, the present meta-analysis does not support the notion that the DRD3 Ser9Gly variant is a genetic risk factor for ET.
Asunto(s)
Temblor Esencial/genética , Predisposición Genética a la Enfermedad , Polimorfismo de Nucleótido Simple , Receptores de Dopamina D3/genética , Genotipo , HumanosRESUMEN
The association between glucocerebrosidase (GBA) mutations and Parkinson's disease (PD) is attracting increased attention worldwide. Results from previous studies on the association of GBA mutations with PD in different ethnicities remain contradictory. In order to derive a more comprehensive understanding of the relationship between the most common GBA mutations, L444P and N370S and PD susceptibility, an updated meta-analysis was performed by searching PUBMED, EMBASE, MEDLINE, and EBSCO databases. Twenty five studies including 9, 599 cases and 13, 541 controls were collected in the end. The summary of odds ratios (OR) and corresponding 95% confidence intervals (CI) were estimated using fixed- and random-effects models, when appropriate. Overall, our meta-analysis provided evidence that both were risk factors associated with increased PD susceptibility. When stratified by ethnicities, the associations varied among different ethnical origins.
Asunto(s)
Predisposición Genética a la Enfermedad/genética , Glucosilceramidasa/genética , Enfermedad de Parkinson/genética , Estudios de Casos y Controles , Etnicidad/genética , Humanos , Mutación/genéticaRESUMEN
It is well known that neural stem cells (NSC) could promote the repairment after spinal cord injury, but the underlying mechanism remains to be elucidated. This study showed that the transplantation of NSC significantly improved hindlimb locomotor functions in adult rats subjected to transection of the spinal cord. Biotin dextran amine tracing together with the stimulus experiment in motor sensory area showed that little CST regeneration existed and functional synaptic formation in the injury site. Immunocytochemistry and RT-PCR demonstrated the secretion of NGF, BDNF, and NT-3 by NSC in vitro and in vivo, respectively. However, only mRNA expression of BDNF and NT-3 but not NGF in injury segment following NSC transplantation was upregulated remarkably, while caspase-3, a crucial apoptosis gene, was downregulated simultaneously. These provided us a clue that the functional recovery was correlated with the regulation of BDNF, NT-3, and caspase-3 in spinal cord transected rats following NSC transplantation.