RESUMEN
Reactive oxygen species (ROS) including superoxide (O2â¢-) play an important role in a variety of diseases, including Alzheimer's Disease, cancer, and atherosclerosis. Early reports showed that O2â¢- is a stimulant for collagen synthesis. However, the mechanism remains incompletely understood. Here we showed that LY83583 (6-anilinoquinoline-5,8-quinone), a substance known to induce O2â¢- production by smooth muscle cell (SMC), increases Type I collagen secretion. This effect could be blocked by treating the cells with Tiron, a scavenger for O2â¢-. LY83583-induced Type I collagen secretion required P4HA1 and P4HA2. Knockout of either P4ha1 or P4ha2 greatly reduced LY83583-stimulated Type I collagen maturation whereas silencing of both P4ha1 and P4ha2 completely blocked LY83583-induced Type I collagen maturation. Although significantly more hydroxyproline on purified Type I collagen was detected from LY83583 treated mouse embryonic fibroblast (MEF) cells by mass spectrometry, the level of prolyl 4-hydroxylases was not altered. Thus, LY83583 might increase the enzymatic activity of prolyl 4-hydroxylases to increase Type I collagen maturation. In addition, we found that LY83583 activated prolyl 4-hydrolases differed from ascorbate-activated prolyl 4-hydroxylase in two aspects: (1) LY83583 activated both P4HA1 and P4HA2 involved in collagen maturation whereas ascorbate mainly stimulated P4HA1 in collagen maturation; (2) LY83583 did not induce N259 glycosylation on P4HA1 as ascorbate did. The mechanisms remain to be investigated.
Asunto(s)
Colágeno Tipo I/metabolismo , Prolil Hidroxilasas/metabolismo , Superóxidos/metabolismo , Aminoquinolinas/farmacología , Animales , Aorta/citología , Colágeno Tipo I/genética , Embrión de Mamíferos/citología , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Glicosilación , Hidroxilación , Ratones , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Péptidos/metabolismo , Prolina/metabolismo , Transcripción Genética/efectos de los fármacosRESUMEN
Alzheimer's disease (AD), one of the most common types of chronic neurodegenerative diseases, is pathologically characterized by the formation of amyloid ß (Aß) peptidecontaining plaques and neurofibrillary tangles. Among Aß peptides, Aß142 induces neuronal toxicity and neurodegeneration. In our previous studies, Cdk5 was found to regulate Aß142induced mitochondrial fission via the phosphorylation of dynaminrelated protein 1 (Drp1) at Ser579. However, whether blockage of Drp1 phosphorylation at Ser579 protects neurons against Aß142induced degeneration remains to be elucidated. Thus, the aim the present study was to examine the effect of mutant Drp1S579A on neurodegeneration and its underlying mechanism. First, the phosphorylationdefect (phosphodefect) mutant, LentiDrp1S579A was constructed. Phosphodefect Drp1S579A expression was detected in primary cultures of mouse cortical neurons infected with LentiDrp1S579A using western blotting and it was found to successfully attenuate the phosphorylation of endogenous Drp1 at Ser579. In primary neuronal cultures, the neuronal processes were evaluated under microscopy. Treatment with 10 µM Aß142 significantly decreased dendritic density and length, spine outgrowth and synapse number. As expected, infection of neurons with LentiDrp1S579A efficiently alleviated the inhibitory effect of Aß142 on neurite outgrowth and synapse density. In addition, infection with LentiDrp1S579A abolished the cleavage of caspase3 and apoptosis in neurons exposed to Aß142. Thus, the current data demonstrated that blockage of Drp1 phosphorylation at Ser579 may be an effective strategy to protect neurons against Aß142induced degeneration and apoptosis. These findings underline the therapeutic potential of targeting Drp1 in the treatment of AD.