Oleic acid ameliorates palmitic acid induced hepatocellular lipotoxicity by inhibition of ER stress and pyroptosis

Abstract

Background: Pyroptosis is a novel programmed cell death. It is identified as caspase-1 dependent and characterized by plasma-membrane rupture and release of proinflammatory intracellular contents inculuding IL-1 beta and IL-18. Pyroptosis is distinct from other forms of cell death, especially apoptosis that is characterized by nuclear and cytoplasmic condensation and is elicited via activation of a caspase cascade. In pyroptosis, gasdermin D (GSDMD) acts as a major executor, while NLRP3 related inflammasome is closely linked to caspase-1 activation. Given that pyroptosis has played a critical role in the progression of non-alcoholic steatohepatitis (NASH), here, we investigated whether the regulation of pyroptosis activation is responsible for the protective role of monounsaturated oleic acids in the context of hepatocellular lipotoxicity. Methods: Human hepatoma cell line HepG2 cells were exposed to palmitic acid (PA) with or without oleic acids (OA) or/and endoplasmic reticulum (ER) stress inhibitor tauroursodeoxycholic acid (TUDCA) for 24 h. Besides, the cells were treated with the chemical ER stressor tunicamycin (TM) with or without OA for 24 h as well. The expressions of pyroptosis and ER stress related genes or proteins were determined by real-time PCR, Western blot or immunofluorescence. The morphology of pyroptosis was detected by acridine orange and ethidium bromide (AO/EB) staining. The release of IL-1 beta and tumor necrosis factor alpha (TNF-α) was determined by ELISA. Sprague-Dawley (SD) rats were fed with high fat diet (HFD) for 16 w, then, HFD was half replaced by olive oil to observe the protective effects of olive oil. The blood chemistry were analyzed, and the liver histology and the expressions of related genes and proteins were determined in the liver tissues. Results: We demonstrated that PA impaired the cell viability and disturbed the lipid metabolism of HepG2 cells (P < 0.01), but OA robustly rescued cells from cell death (P < 0.001). More importantly, we found that instead of cell apoptosis, PA induced significant pyroptosis, evidenced by remarkably increased mRNA and protein expressions of inflammasome marker NLRP3, Caspase-1 and IL-1beta, as well as cell membrane perforation driving protein GSDMD (P < 0.05). Furthermore, we demonstrated that the PA stimulated ER stress was causally related to pyroptosis. The enhanced expressions of ER stress markers CHOP and BIP were found subcellular co-located to pyroptosis markers NLRP3 and ASC. Additionally,TM was able to induce pyroptosis like PA did, and ER stress inhibitor TUDCA was able to inhibit both PA and TM induced ER stress as well as pyroptosis. Furthermore, we demonstrated that OA substantially alleviated either PA or TM induced ER stress and pyroptosis in HepG2 cells (P < 0.01). In vivo, only olive oil supplementation did not cause significant toxicity, while HFD for 32 w obviously induced liver steatosis and inflammation in SD rats (P < 0.05). Half

The NLRP3 Inflammasome: Metabolic Regulation and Contribution to Inflammaging

“…In addition to activation of the NLRP3 inflammasome, palmitic acid has also been shown to cause many cellular perturbations such as ER stress, decreased cellular viability, loss of mitochondrial membrane potential, and the induction of many inflammatory genes such as IL-6 and TNF [79,80]. Mechanistically, palmitic acid and other saturated fatty acids can be crystallized in macrophages, which causes lysosomal membrane rupture, a known NLRP3 inflammasome trigger [76].…”

“…In contrast, oleic acid inhibits and even mitigates the effects of palmitic acid, such that the treatment of macrophages with oleic acid inhibits IL-1β release in a dose-dependent manner [76]. Additionally, in HepG2 cells, oleic acid is able to decrease NLRP3 inflammasome activation and pyroptosis by reducing ER stress to counteract the effects of palmitic acid [80].…”

Regulating metabolic inflammation by nutritional modulation

“…50 Regarding liver function, administration of oleic acid has been shown to inhibit ER stress, alleviating hepatocellular lipotoxicity. 51 Similarly, inhibiting IRE1a-mediated inflammasome activation prevents the progression of NAFLD to steatohepatitis. 52 These discoveries paved the way to establishment of drugmediated therapies targeting the ER stress response (caspase-1, IRE1a, or acetyl-coenzyme A carboxylase inhibitors) to slow down the development of liver disease, and despite the fact that hypernutrition concurs with ER stress to promote hepatic tumorigenesis, 53 therapeutic nutritional approaches remain to be developped in this regard.…”

“…115 In the liver, replacement of SFAs with MUFAs reduced hepatic GSDMD/-N, caspase-1, and p20 levels, indicating attenuation of NLRP3 activation. 51 Furthermore, in vivo saturated fat diet displays increased hepatic inflammation with increased serum amyloid A1 (SAA1) protein expression paired with reduced cholesterol transport compared with the low-fat diet control but not with monounsaturated fat diet. 87 In humans, a diet high in MUFAs decreases liver fat in both prediabetes and T2D compared with an isocaloric highcarbohydrate and high-fiber diet.…”

Hesperidin and Myricetin Attenuated Non-Alcoholic Fatty Liver Disease (NAFLD) in HepG2 Cells

“…Our study found that both PA and OA induced NAFLD via induction of PPARs, SREBP, ACC, ACOX, and FAS, which corresponds with previous reports that PA and OA disturbed lipid metabolism and induced NAFLD in HepG2 cells. 28,29 In one study, PA impaired cell viability and disturbed lipid metabolism in HepG2 cells, but OA robustly rescued cells from cell death by inhibition of endoplasmic reticulum stress and pyroptosis. 28 Moreover, in another study, OA reduced PA-induced oxidative stress, endoplasmic reticulum stress, and mitochondrial dysfunction in rat hepatocytes.…”