IN VITRO STUDY TO ASSESS THE POTENTIAL OF SHORT CHAIN FATTY ACIDS (SCFA) AS THERAPEUTIC AGENTS FOR ALZHEIMER'S DISEASE

Abstract

A diet rich in fibre has components which are possibly advantageous for improved cognitive performance. Such a diet is characterised by high consumption of vegetables, legumes, fruits, and cereals and unsaturated fatty acids; evidence suggests that diets rich in fibre reduce the associated risk factors of AD via formation of short chain fatty acids (SCFAs). SCFAs are 2-carbon (C2) to 5-carbon (C5) weak acids; mainly acetate (C2), propionate (C3), and butyrate (C4). Our understanding of how fibre intake might influence biomarkers (tau/ Aβ) of Alzheimer's disease (AD) is limited. This project was aimed to address this significant knowledge gap by examining the capacity of SCFAs to modify the levels of Aβ at cellular level. Human hepatocellular liver carcinoma cells (HepG2 cells) were treated with SCFAs and Aβ. Cells were treated with different concentrations of Aβ and SCFA (Acetate, butyrate, propionate) for 48 hours. Western blotting, THT, MTS and AFM was carried out. HepG2 cells cells were cultured in Dulbecco's modified Eagle's medium with Glutamax, fetal calf serum and penStrep. Cells were grown in the 75ml flasks to 70% confluence. After changing the medium cells were treated with different concentration of SCFA ranging from 1-10 mM for 24 hours. Then cells were treated again with respective compounds and incubated with 1 μ M Aβ. Abeta was prepared as homogeneous and as unaggregated and as oligomers. Direct effect of SCFA and Aβ was analysed using THT and AFM, cell toxicity was measured using MTS and LDH assays, cell lysates and supernatants were analysed by western immunoblotting. A significant decrease in Aβ levels and Aβ aggregation was observed by Western blot and ThT assay respectively, in cells treated with SCFAs compared to cells without SCFA treatment: Aβ levels measured in cell lysate and medium were reduced by 60-70%. This reduction in Aβ levels was most pronounced with butyrate treatment. ThT and AFM images showed the reduction of aggregation. Cellular morphology was also monitored by microscopy each day and cells appeared healthier after treatment with SCFA and amyloid beta for 48 hours. This observation was supported by the results of MTS and LDH cell viability assays. The results of our in vitro experiments suggest that SCFAs have a potential to reduce Aβ levels in cells. Further, inhibition of the aggregation of Aβ indicates the potential of reduction of Aβ-mediated toxicity under in vitro conditions. The next step is to confirm these results in an in vivo setting using an animal model of AD. Therefore, a small pilot study using a transgenic mouse model of AD (5XFAD) will be carried out.