RESUMO
The most recent research concerning amyotrophic lateral sclerosis (ALS) emphasizes the role of glia in disease development. Thus, one can suspect that the effective therapeutic strategy in treatment of ALS would be replacement of defective glia. One of the basic problems with human glial progenitors (hGRPs) replacement strategies is the time needed for the cells to become fully functional in vivo. The lifespan of most popular high copy number SOD1 mutant mice might be too short to acknowledge benefits of transplanted cells. We focused on developing immunodeficient rag2-/- model of ALS with lower number of transgene copies and longer lifespan. The obtained hSOD1/rag2 double mutant mice have been characterized. QPCR analysis revealed that copy number of hSOD1 transgene varied in our colony (4-8 copies). The difference in transgene copy number may be translated to significant impact on the lifespan. The death of long- and short-living hSOD1/rag2 mice is preceded by muscular weakness as early as one month before death. Importantly, based on magnetic resonance imaging we identified that mutant mice demonstrated abnormalities within the medullar motor nuclei. To conclude, we developed long-living double mutant hSOD1/rag2 mice, which could be a promising model for testing therapeutic utility of human stem cells.
Assuntos
Esclerose Lateral Amiotrófica/diagnóstico por imagem , Variações do Número de Cópias de DNA , Proteínas de Ligação a DNA/genética , Superóxido Dismutase-1/genética , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/imunologia , Esclerose Lateral Amiotrófica/fisiopatologia , Animais , Proteínas de Ligação a DNA/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Feminino , Técnicas de Inativação de Genes , Humanos , Hospedeiro Imunocomprometido , Masculino , Camundongos , Camundongos Transgênicos , Dobramento de Proteína , Índice de Gravidade de Doença , Medula Espinal/diagnóstico por imagem , Medula Espinal/metabolismo , Superóxido Dismutase-1/química , Superóxido Dismutase-1/metabolismo , Núcleo Motor do Nervo Trigêmeo/diagnóstico por imagem , Núcleo Motor do Nervo Trigêmeo/metabolismoRESUMO
The neurons in the trigeminal mesencephalic nucleus (Vmes) innervate jaw-closing muscle spindles and periodontal ligaments, and play a crucial role in the regulation of jaw movements. Recently, it was shown that many boutons that form synapses on them are immunopositive for glycine (Gly+), suggesting that these neurons receive glycinergic input. Information about the glycine receptors that mediate this input is needed to help understand the role of glycine in controlling Vmes neuron excitability. For this, we investigated the expression of glycine receptor subunit alpha 3 (GlyRα3) and gephyrin in neurons in Vmes and the trigeminal motor nucleus (Vmo), and the Gly+ boutons that contact them by light- and electron-microscopic immunocytochemistry and quantitative ultrastructural analysis. The somata of the Vmes neurons were immunostained for GlyRα3, but not gephyrin, indicating expression of homomeric GlyR. The immunostaining for GlyRα3 was localized away from the synapses in the Vmes neuron somata, in contrast to the Vmo neurons, where the staining for GlyRα3 and gephyrin were localized at the subsynaptic zones in somata and dendrites. Additionally, the ultrastructural determinants of synaptic strength, bouton volume, mitochondrial volume, and active zone area, were significantly smaller in Gly+ boutons on the Vmes neurons than in those on the Vmo neurons. These findings support the notion that the Vmes neurons receive glycinergic input via putative extrasynaptic homomeric glycine receptors, likely mediating a slow, tonic modulation of the Vmes neuron excitability.