A Systems Approach to ER Stress Pathway Dynamics Reveals the Impact of Freeze-Dried Broccoli Extract to Mitigate Inflammation in Human Adipocytes through the Mevalonate Pathway

Abstract

Background: Delivery of nutrient excess in obesity causes endoplasmic reticulum (ER) stress within adipose tissue; this activates the unfolded protein response (UPR) and contributes to type 2 diabetes mellitus (T2DM) risk. This study aimed to utilise freeze-dried broccoli extract (BE) as a nutrient to mitigate such cellular damage in human adipocytes, and create a mathematical model to understand pathway dynamics. Methods: Differentiated human adipocytes (Chub-S7; n=6) were treated with BE (10ng/ml) alone or combined with tunicamycin (Tun; 750ng/ml), an inducer of ER stress. UPR proteins (BiP, PERK, P-PERK, eIF2α, P-eIF2α) were measured (18 time points; 0hr-72hr) using Western Blot and transcriptomics. Mass action kinetics was used to create ordinary differential equations (ODEs) to model the UPR for predictive analysis. Results: Tun increased UPR proteins 9.5 fold (P<0.05), whilst BE+Tun reduced ER stress proteins by 94% (P<0.05). Transcriptomic analysis highlighted significant changes in the mevalonate pathway with BE (P<0.05), whilst time series data identified oscillatory behaviour of UPR proteins. Finally, modelling pathway dynamics, time-series analysis improved the error between model output and experimental data by 23%, yielding a new enhanced qualitative model. Conclusion: BE acts to alleviate ER stress in human adipocytes by reducing the UPR through the mevalonate pathway. Furthermore, modelling pathway dynamics using experimental data may provide insight into predicting nutrient capabilities to reduce inflammation. Disclosure A. Murphy: None. S. Azharian: None. G. Tripathi: None. G. Barker: None. M.J. Chappell: None. P.G. McTernan: None.