RESUMO
Niemann-Pick type C disease (NPC) is a neurodegenerative genetic disorder caused by accumulation of lipids, especially cholesterol, in the perinuclear space. U18666A is a cholesterol transport-inhibiting agent, being used to mimic NPC, mainly in fibroblasts. The objective of this study was to observe the effect of the drug U18666A, which causes the accumulation of cholesterol in the cytoplasm of astrocytes from newborn rats, on some lysosomal hydrolase activities. Filipin staining and fluorescence microscopy, through CellM software, were used for visualization and quantification of cholesterol. The dose of U18666A that provided the greatest accumulation of cholesterol was that of 0.25 µg/mL in incubation for 48 h. Primary rat astrocytes incubated with the drug (NPC) showed a significantly higher amount of cholesterol than those without U18666A (controls). The measurement of activity of enzymes sphingomyelinase and beta-glucosidase in astrocytes of rats with NPC was significantly lower than that of control astrocytes, which is consistent with the disease in humans. The activity of the enzyme beta-galactosidase showed no significant difference between both groups. We concluded that U18666A appears to be an excellent intracellular cholesterol transport-inhibiting agent affecting some metabolic pathways in astrocytes of young rats, which mimics NPC in these animals. Just like the change in the activity of lysosomal enzymes has been demonstrated, other biochemical parameters of the cell can be tested with this animal model, thus contributing to a better understanding of the disease.
Assuntos
Androstenos/farmacologia , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Esfingomielina Fosfodiesterase/metabolismo , beta-Galactosidase/metabolismo , beta-Glucosidase/metabolismo , Animais , Anticolesterolemiantes/farmacologia , Células Cultivadas , Colesterol/metabolismo , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Hidrolases/metabolismo , Espaço Intracelular/metabolismo , Lisossomos/metabolismo , RatosRESUMO
This study investigates the miniaturization of the screening technique using dried blood spots on filter paper (DBS) to measure GBA and CT activities, and GBA and ß-galactosidase activities in leukocytes. 274 DBS from individuals with suspected GD were screened for 1.5 years. Of these, we confirmed the diagnosis in 13.5%. The miniaturization of the DBS and leukocyte techniques afforded to reduce costs and sample size appropriate for a reliable diagnosis.
