Screening and identification of differentially expressed genes in goose hepatocytes exposed to free fatty acid

Abstract

The overaccumulation of triglycerides in hepatocytes induces hepatic steatosis; however, little is known about the mechanism of goose hepatic steatosis. The aim of this study was to define an experimental model of hepatocellular steatosis with TG overaccumulation and minimal cytotoxicity, using a mixture of various proportions of oleate and palmitate free fatty acids (FFAs) to induce fat-overloading, then using suppressive subtractive hybridization and a quantitative PCR approach to identify genes with higher or lower expression levels after the treatment of cells with FFA mixtures. Overall, 502 differentially expressed clones, representing 21 novel genes and 87 known genes, were detected by SSH. Based on functional clustering, up- and down-regulated genes were mostly related to carbohydrate and lipid metabolism, enzyme activity and signal transduction. The expression of 20 selected clones involved with carbohydrate and lipid metabolism pathways was further studied by quantitative PCR. The data indicated that six clones similar to the genes ChREBP, FoxO1, apoB, IHPK2, KIF1B, and FSP27, which participate in de novo synthesis of fatty acid and secretion of very low density lipoproteins, had significantly lower expression levels in the hepatocytes treated with FFA mixtures. Meanwhile, 13 clones similar to the genes DGAT-1, ACSL1, DHRS7, PPARα, L-FABP, DGAT-2, PCK, ACSL3, CPT-1, A-FABP, PPARβ, MAT, and ALDOB had significantly higher expression levels in the hepatocytes treated with FFA mixtures. These results suggest that several metabolic pathways are altered in goose hepatocytes, which may be useful for further research into the molecular mechanism of goose hepatic steatosis.

Chromosome-level genome and population genomics reveal evolutionary characteristics and conservation status of Chinese indigenous geese

“…Positive selection analysis identified a series of PSGs enriched in lipid metabolism and RNA processing. Involved genes such as APOB participate in de novo synthesis of fatty acids and were associated with goose hepatic steatosis 38 . Cholesteryl ester transfer protein ( CETP ) has been shown to play a role in liver lipid metabolism 39 .…”

Fads1 and 2 are promoted to meet instant need for long-chain polyunsaturated fatty acids in goose fatty liver

“…In this study, goose primary hepatocytes were treated with higher dosages (25 or 50 mM) of glucose due to the following consideration: (1) The acute effect of strong stimulation (high glucose) may better mimic the chronic effect of hyperglycemia on the hepatocytes in goose liver than that of weak stimulation (low glucose as in blood); (2) when insulin resistance occurs in goose fatty liver, glucose transporter may pump a large amount of glucose into the hepatocytes in the liver in addition to the diffusion of glucose, which may lead to intracellular glucose level much higher than blood glucose level; (3) Treatment with high level of glucose was previously reported by some in vitro studies [55]. In regard to treatments with 0.25/0.5 mM fatty acids and 100/200 nM insulin, previous studies were also referenced, including those showing serum-free fatty acids were above 0.5 mM in mammalian animals [56][57][58], as well as those using the same or even higher dosages for in vitro studies with mammalian cells and avian cells [55,59,60]. These in vitro studies indicated that the expression of Fads genes was differentially regulated by these factors.…”

Vitamin D/VDR signaling suppresses microRNA‐802‐induced apoptosis of keratinocytes in oral lichen planus

“…Caspase-3 activity assay HOK cells were lysed by using a lysis buffer (1% Triton X-100, 1 mM DTT, 25 mM Hepes, 115 mM NaCl, 1 mM KH 2 PO 4 , 4 mM KCl, and 1 ml/ml protease inhibitor cocktail) on ice for 10 min, then collected and centrifuged to have the supernatants as cell lysate. The caspase-3 activity in cell lysates was evaluated by using a specific fluorogenic caspase substrate Ac-DEVD-AFC (BioVision, Milpitas, CA, USA) and measured at Ex.360/Em.530 on a plate reader as previously described (33,34).…”