RESUMO
Aging causes major alterations of all components of the neurovascular unit and compromises brain blood supply. Here, we tested how aging affects vascular reactivity in basilar arteries from young (<10 weeks; y-BA), old (>22 months; o-BA) and old (>22 months) heterozygous MYPT1-T-696A/+ knock-in mice. In isometrically mounted o-BA, media thickness was increased by â¼10% while the passive length tension relations were not altered. Endothelial denudation or pan-NOS inhibition (100 µmol/L L-NAME) increased the basal tone by 11% in y-BA and 23% in o-BA, while inhibition of nNOS (1 µmol/L L-NPA) induced â¼10% increase in both ages. eNOS expression was â¼2-fold higher in o-BA. In o-BA, U46619-induced force was augmented (pEC50 â¼6.9 vs. pEC50 â¼6.5) while responsiveness to DEA-NONOate, electrical field stimulation or nicotine was decreased. Basal phosphorylation of MLC20-S19 and MYPT1-T-853 was higher in o-BA and was reversed by apocynin. Furthermore, permeabilized o-BA showed enhanced Ca2+-sensitivity. Old T-696A/+ BA displayed a reduced phosphorylation of MYPT1-T696 and MLC20, a lower basal tone in response to L-NAME and a reduced eNOS expression. The results indicate that the vascular hypercontractility found in o-BA is mediated by inhibition of MLCP and is partially compensated by an upregulation of endothelial NO release.
Assuntos
Acetofenonas/farmacologia , Envelhecimento , Artéria Basilar/fisiologia , Músculo Liso Vascular/fisiologia , Fosfatase de Miosina-de-Cadeia-Leve/antagonistas & inibidores , Óxido Nítrico Sintase Tipo III/metabolismo , Animais , Inibidores Enzimáticos , Camundongos , Fosfatase de Miosina-de-Cadeia-Leve/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Fosforilação , Subunidades Proteicas/metabolismo , VasoconstriçãoRESUMO
BACKGROUND: Recent progress in the cardiotypic differentiation of embryonic and somatic stem cells opens novel prospects for the treatment of cardiovascular disorders. The aim of the present study was to develop a novel surgical approach that allows standardized cellular cardiomyoplasty in mouse with low-perioperative mortality. METHODS: Reproducible transmural lesions were generated by cryoinjury followed by intramural injection of embryonic cardiomyocytes using a newly designed holding device and vital dye staining. This approach was validated with a transgenic mouse model, in which the live reporter gene-enhanced green fluorescent protein (EGFP) is under control of a cardiac-specific promoter. RESULTS: The perioperative mortality was 10%. The engrafted EGFP-positive cardiomyocytes could be identified in a high percentage (72.2%, n=36) of operated animals. CONCLUSIONS: This novel approach enables reliable cellular replacement therapy in mouse and greatly facilitates the analysis of its molecular, cellular, and functional efficacy.