Regulation of proliferation rates in human astrocytes by modulating mitochondrial and cell metabolic activity.

Abstract

Astrocytes contribute to the normal function and maintenance of the health of the central nervous system (CNS). The abnormal behavior of astrocytes can lead to a major alteration in neuronal function contributing to the pathogenesis of several neurological diseases, including Alzheimer's disease (AD). Dysfunctional astrocytes have been linked to the onset and development of overreacting inflammation in CNS tissue which is a major contributing factor to the onset and progression of AD. We hypothesize that by modulating mitochondrial activity and cell metabolism using antioxidants, key characteristics of astrogliosis could be regulated. Mainly, we believe that some antioxidants can help to regulate astrocyte response and promote their return back to a quiescent state. In this work, we explored the use of two antioxidant molecules (A1 and A2) as a way to regulate the metabolic activity, proliferation, and overreactive behavior of glial cells. Human astrocytes were cultured for five days in growth media supplemented with 0, 25, or 50 μM A1 or A2. Changes in DNA levels were used to calculate the doubling time of the cell population. DNA levels were quantified using PicoGreen dsDNA quantitation assay. Astrocytes were stained with JC-1 and fluorescence readings were taken to evaluate mitochondrial activity. Changes in metabolic activity were evaluated using the Vibrant Metabolic Assay and Adenosine Triphosphate (ATP) production. Quantitative assessments demonstrate that increasing levels of A1 significantly reduce human astrocyte proliferation while the addition of A2 at the chosen concentrations does not appear to have a significant effect on cell proliferation. The preliminary data for ATP production and metabolic activity illustrates that A1 appears to decrease cell metabolic activity which may lead to lower cell proliferation. In addition, quantitative analysis of the JC-1 mitochondrial staining suggests that the addition of A1 does not significantly impact mitochondrial membrane potential while increasing levels of A2 enhanced the polarization of the mitochondrial membrane. The modulation of mitochondrial and metabolic activity using antioxidant molecules may be used as a viable strategy to regulate the overreactive proliferative response of human astrocytes during astrogliosis.