Hydrolysis products generated by Lipoprotein Lipase and their association with Oxidative Stress in THP-1 Macrophages

Marzana Monefa
MSc. Student
Department of Biochemistry

Abstract:

Lipoprotein lipase (LPL) is an extracellular lipase that hydrolyzes triglycerides from triglyceride-rich lipoproteins (Lp) within the bloodstream. In the arteries, LPL expression has been observed within macrophages that negatively influence atherosclerosis and promote cardiovascular disease. Previously, our lab reported that Lp hydrolysis products generated by LPL resulted in the upregulated expression of 63 small nucleolar RNAs (snoRNA) within a human macrophage model. Interestingly, the hydrolysis of very-low-density lipoprotein (VLDL) by LPL induces reactive oxygen species (ROS) production in human aortic endothelial cells, and cell stress induces snoRNA expression in various cell models; the expression can be impeded by inhibiting NADPH oxidase (Nox). The project aims to assess the role of LPL in oxidative stress when it hydrolyzes human Lp using in vitro and in silico approaches. For in vitro studies, human VLDL and chylomicron hydrolysis products 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 using a ROS assay and thiobarbituric acid reactive substances (TBARS) assay, respectively. The ROS levels within media and cells increased in response to increasing levels of hydrolysis products of both Lps by LPL. However, the resultant media and cellular ROS decreased with the addition of the NOXi treatment as expected. Additionally, media malondialdehyde (MDA) demonstrated reduction, although the cellular MDA displayed increment when Noxi was treated to the Lp hydrolysis products by LPL. For In silico analysis, available transcriptomic datasets for Lp lipolysis by LPL in two cell lines showed an upregulation of stress-inducing genes. Future work will include examining the THP-1 cells for endoplasmic reticulum stress and RNA integrity with or without Nox inhibitors to confirm if Nox is an effector of Lp hydrolysis product-mediated snoRNA expression.