RESUMO
Neurotensin (NTS)-polyplex is a multicomponent nonviral vector that enables gene delivery via internalization of the neurotensin type 1 receptor (NTSR1) to dopaminergic neurons and cancer cells. An approach to improving its therapeutic safety is replacing the viral karyophilic component (peptide KPSV40; MAPTKRKGSCPGAAPNKPK), which performs the nuclear import activity, by a shorter synthetic peptide (KPRa; KMAPKKRK). We explored this issue and the mechanism of plasmid DNA translocation through the expression of the green fluorescent protein or red fluorescent protein fused with KPRa and internalization assays and whole-cell patch-clamp configuration experiments in a single cell together with importin α/ß pathway blockers. We showed that KPRa electrostatically bound to plasmid DNA increased the transgene expression compared with KPSV40 and enabled nuclear translocation of KPRa-fused red fluorescent proteins and plasmid DNA. Such translocation was blocked with ivermectin or mifepristone, suggesting importin α/ß pathway mediation. KPRa also enabled NTS-polyplex-mediated expression of reporter or physiological genes such as human mesencephalic-derived neurotrophic factor (hMANF) in dopaminergic neurons in vivo. KPRa is a synthetic monopartite peptide that showed nuclear import activity in NTS-polyplex vector-mediated gene delivery. KPRa could also improve the transfection of other nonviral vectors used in gene therapy.
Assuntos
Portadores de Fármacos/síntese química , Técnicas de Transferência de Genes , Vetores Genéticos/administração & dosagem , Neurotensina/administração & dosagem , Fragmentos de Peptídeos/síntese química , Transporte Ativo do Núcleo Celular , Animais , Linhagem Celular , Núcleo Celular/metabolismo , Neurônios Dopaminérgicos/citologia , Neurônios Dopaminérgicos/metabolismo , Terapia Genética/métodos , Vetores Genéticos/genética , Masculino , Camundongos , Modelos Animais , Nanopartículas/química , Neurotensina/genética , Neurotensina/farmacocinética , Técnicas de Patch-Clamp , Plasmídeos/genética , Ratos , Receptores de Neurotensina/metabolismo , Análise de Célula Única , Técnicas EstereotáxicasRESUMO
Se describe el caso de un varón de 7 años que se presenta clínicamente con episodios recidivantes de fiebre, aftas y manifestaciones de meningoencefalitis, con hiperproteinorraquia y pleocitosis linfocitaria. Se descartaron otros diagnósticos diferenciales. Con imágenes patológicas en RNM de iso o hipointensidad en T1, hiperintensidad en T2 y Flair, en mesencéfalo, pedúnculos cerebelosos, protuberancia, cuerpo calloso, ganglios basales y cápsula interna y hemisferios cerebrales. En la evolución agrega signos cutáneos cumpliendo con los criterios diagnósticos establecido Grupo Internacional de Estudio de Enfermedad de Behçet. Se realizó tratamiento corticoideo e munosupresor.Destacamos que los criterios diagnósticos internacionales tienen un valor relativo en la etapa prepuberal, dado que las manifestaciones oculares y genitales son infrecuentes y el compromiso parenquimatoso neurológico puede ser su forma de expresión clínica principal.
The case of a 7-year-old Uruguayan boy with recurrent episodes of fever, oral aphthas lesions and meningoencephalitis with elevated protein levels and lymphocytic pleocytosis is presented. Other diagnoses were excluded. MRI showed in acute stages CNS involvement in iso- or hipointense in T1- weighted images and hyperintense in T2-weighted or fluid- attenuated inversion recovery (FLAIR) images in mesodiencephalic junction, cerebellar peduncles, pons, corpus callosum, basal ganglia and internal capsule, and cerebral hemispheres. In the evolution skin lesions appeared, fulfilling the International Study Group Criteria for the Diagnosis of Behçet' Disease. Corticosteroids and immunosuppressant treatment was done.The main difficulty in children is having all the criteria for diagnosis; especially when recurrent genital ulceration and eye lesion rarely appear at this age.
Assuntos
Humanos , Masculino , Criança , Metilprednisolona/uso terapêutico , Síndrome de Behçet/complicações , Síndrome de Behçet/diagnóstico , Síndrome de Behçet/tratamento farmacológico , Encefalopatias/etiologia , Estomatite Aftosa/etiologia , Febre/etiologia , Ressonância Magnética Nuclear BiomolecularRESUMO
Se describe el caso de un varón de 7 años que se presenta clínicamente con episodios recidivantes de fiebre, aftas y manifestaciones de meningoencefalitis, con hiperproteinorraquia y pleocitosis linfocitaria. Se descartaron otros diagnósticos diferenciales. Con imágenes patológicas en RNM de iso o hipointensidad en T1, hiperintensidad en T2 y Flair, en mesencéfalo, pedúnculos cerebelosos, protuberancia, cuerpo calloso, ganglios basales y cápsula interna y hemisferios cerebrales. En la evolución agrega signos cutáneos cumpliendo con los criterios diagnósticos establecido Grupo Internacional de Estudio de Enfermedad de Behçet. Se realizó tratamiento corticoideo e inmunosupresor.Destacamos que los criterios diagnósticos internacionales tienen un valor relativo en la etapa prepuberal, dado que las manifestaciones oculares y genitales son infrecuentes y el compromiso parenquimatoso neurológico puede ser su forma de expresión clínica principal.
The case of a 7-year- old Uruguayan boy with recurrent episodes of fever, oral aphthas lesions and meningoencephalitis with elevated protein levels and lymphocytic pleocytosis is presented. Other diagnoses were excluded. MRI showed in acute stages CNS involvement in iso- or hipointense in T1- weighted images and hyperintense in T2-weighted or fluid- attenuated inversion recovery (FLAIR) images in mesodiencephalic junction, cerebellar peduncles, pons, corpus callosum, basal ganglia and internal capsule, and cerebral hemispheres. In the evolution skin lesions appeared, fulfilling the International Study Group Criteria for the Diagnosis of Behçet' Disease. Corticosteroids and immunosuppressant treatment was done.The main difficulty in children is having all the criteria for diagnosis; especially when recurrent genital ulceration and eye lesion rarely appear at this age.