ABSTRACT
Amyloid beta (Aß) is a peptide cleaved from amyloid precursor protein that contributes to the formation of senile plaques in Alzheimer's disease (AD). The relationship between Aß and astrocyte proliferation in AD remains controversial. Despite pathological findings of increased astrocytic mitosis in AD brains, in vitro studies show an inhibitory effect of Aß on astrocyte proliferation. In this study, we determined the effect of an active fragment of Aß (Aß25-35) on the cell cycle progression of primary rat astrocytes. We found that Aß25-35 (0.3-1.0 µg/ml) enhanced astrocyte proliferation in vitro in a time- and concentration-dependent manner. Increased DNA synthesis by Aß25-35 was observed during the S phase of the astrocyte cell cycle, as indicated by proliferation kinetics and bromodeoxyuridine immunocytochemical staining. Aggregation of Aß25-35 abolished the upregulatory effect of Aß on astrocyte proliferation. Further examination indicated that Aß25-35 affected astrocyte proliferation during early or mid-G1 phase but had no effect on DNA synthesis at the peak of S phase. These results provide insight into the relationship between Aß25-35 and astrocyte cell cycling in AD.
ABSTRACT
Lipofectamine 2000 is a cationic liposome based reagent that provides high transfection efficiency and high levels of transgene expression in a range of mammalian cell types in vitro using a simple protocol. Optimum transfection efficiency and subsequent cell viability depend on a number of experimental variables such as cell density, liposome and DNA concentrations, liposome-DNA complexing time, and the presence or absence of media components such as antibiotics and serum. The importance of these factors in Lipofectamine 2000 mediated transfection will be discussed together with some specific applications: transfection of primary neurons, high throughput transfection, and delivery of small interfering RNAs.
