Hydrolysis products generated by lipoprotein lipase and their association with oxidative stress in THP-1 macrophages

Monefa, Marzana (2022) Hydrolysis products generated by lipoprotein lipase and their association with oxidative stress in THP-1 macrophages. Masters thesis, Memofrial University of Nefoundland.

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Abstract

Lipoprotein lipase (LPL) is an extracellular lipase that hydrolyzes triglycerides (TG) from TG-rich lipoproteins (Lp) within the bloodstream. In the arteries, macrophage LPL expression has been observed to negatively influence atherosclerosis and promote cardiovascular disease. Previously, our laboratory reported that Lp hydrolysis products (HPs) generated by LPL resulted in the upregulated expression of 63 small nucleolar RNAs (snoRNA) within the human macrophage cell. Interestingly, the very low-density lipoprotein (VLDL) HPs by LPL induced the production of reactive oxygen species (ROS), and the cell stress further induced the snoRNA expression in various cell models. The inhibition of NADPH oxidase (NOX) in the cell models diminished snoRNA expression. This project aimed to assess the role of Lp HPs by LPL in oxidative stress using in vitro and in silico approaches. For in vitro studies, human VLDL and chylomicron (CM) HPs generated by LPL were incubated with THP-1 macrophages, with or without a NOX inhibitor (NOXi). The ROS production and lipid peroxidation in the media and cells were analyzed. With an increase in free fatty acid (FFA) concentration from both Lp HPs by LPL, relative to control, there was a decreasing trend in ROS generation for the lowest FFA concentration followed by increasing trend for other FFA concentrations. However, when THP-1 cells were incubated with 0.5 nmol/μL FFA of both Lps HPs by LPL, there was a reduction in ROS with NOXi treatment relative to control. Extracellular ROS was higher with NOXi with 0.5 nmol/μL FFA from VLDL. On the other hand, no change in media malondialdehyde (MDA) with both Lps HPs by LPL was decreased with the addition of NOXi, while the cellular MDA was decreased with or without NOXi. For in silico analyses, available transcriptomic datasets for Lp lipolysis by LPL in two cell lines showed an upregulation of stress-inducing genes and a downregulation of genes associated with cell cycle arrest. Henceforth, my study indicated that Lp HPs by LPL may stimulate foam cell conversion through enhanced NOX-mediated ROS and extracellular MDA levels, which may lead to cellular stress and cell death.

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