Role of Carboxylesterase 1 in THP-1 Human Macrophages Under Basal Conditions or Treated with LPS+IFN Gamma (M1) OR with IL4 (M2)

Macrophages are one of the major immune cell types responsible for maintaining the innate immunity of the host. They can be polarized into one of the two sub populations in vitro. M1 which is bactericidal and pro inflammatory, and M2 which is anti inflammatory and active in tissue repair. Carboxylesterases are members of the serine hydrolase superfamily. They can hydrolyze ester-containing compounds into alcohol- and carboxylic acid-containing products. About six isoforms have been identified in humans, and CES1 and CES2 are the most studied and best characterized. CES1 is a triacylglycerol hydrolase. Triacylglycerols are primarily found in cytoplasmic lipid droplets and are the major form of fat storage in the cells. Hydrolysis of triacylglycerol liberates free fatty acids that are crucial for cellular metabolic activities. Liberated fatty acids such as polyunsaturated fatty acids (PUFA) become substrates for enzymes that produce eicosanoids, while other fatty acids act as ligands for membrane receptors and nuclear receptors, such as toll-like receptor 4 (TLR4) and PPAR gamma respectively. We sought to understand how the inactivation of CES1 in the human THP-1 macrophage cell line would impact lipid metabolism and macrophage phenotype under resting conditions and following stimulation. Control and CES1 knockdown (KD) THP-1 monocytic cells were differentiated into macrophages with 20 nM PMA over 72 h, then rested for 24 h prior to being treated with either 1-ug/mL LPS + 100-ng/mL interferon gamma (M1) or 100-ng/mL IL-4 (M2) for 24 h, or they were left untreated (M0) for the same length of time. Total RNA was extracted using QIAshredder (QIAGEN) and sent to Novogene Inc (Sacramento, CA) for library construction, RNA sequencing and data processing. Our data present a transcriptomic snapshot when CES1 is knocked down in THP-1 macrophages during the resting state and polarized state. Overall design: To investigate the role of cabroxylestesrase 1 in naïve (M0), M1, and M2 THP-1 macrophages Carboxylesterase 1 gene was stably knocked down with shRNA using a lentivirus system, and its level of expression was verified to be at least 85% reduced. We perfomed comparative gene expression profiling of the control and knocked down (CES1KD) macrophages using three experimental groups. Experimental groups were defined as follows: resting (M0), LPS + interferon gamma (M1), and IL4 (M2), each analyzed in quadruplicate.