Docosahexaenoic acid enhances hepatic serum amyloid A expression via protein kinase A-dependent mechanism.

Serum amyloid A (SAA) reduces fat deposition in adipocytes and hepatoma cells. Human SAA1 mRNA is increased by docosahexaenoic acid (DHA) treatment in human cells. These studies asked whether DHA decreases fat deposition through SAA1 and explored the mechanisms involved. We demonstrated that DHA increased human SAA1 and C/EBPbeta mRNA expression in human hepatoma cells, SK-HEP-1. Utilizing a promoter deletion assay, we found that a CCAAT/enhancer-binding protein beta (C/EBPbeta)-binding site in the SAA1 promoter region between -242 and -102 bp was critical for DHA-mediated SAA1 expression. Mutation of the putative C/EBPbeta-binding site suppressed the DHA-induced SAA1 promoter activity. The addition of the protein kinase A inhibitor H89 negated the DHA-induced increase in C/EBPbeta protein expression. The up-regulation of SAA1 mRNA and protein by DHA was also inhibited by H89. We also demonstrated that DHA increased protein kinase A (PKA) activities. These data suggest that C/EBPbeta is involved in the DHA-regulated increase in SAA1 expression via PKA-dependent mechanisms. Furthermore, the suppressive effect of DHA on triacylglycerol accumulation was abolished by H89 in SK-HEP-1 cells and adipocytes, indicating that DHA also reduces lipid accumulation via PKA. The observation of increased SAA1 expression coupled with reduced fat accumulation mediated by DHA via PKA suggests that SAA1 is involved in DHA-induced triacylglycerol breakdown. These findings provide new insights into the complicated regulatory network in DHA-mediated lipid metabolism and are useful in developing new approaches to reduce body fat deposition and fatty liver.

The effect of feed restriction on expression of hepatic lipogenic genes in broiler chickens and the function of SREBP1.

To study the role of sterol regulatory element-binding proteins (SREBP) in lipogenesis and cholesterol synthesis in the chicken, two experiments were carried out. In the first study, seven-week-old broilers (n=16) were allocated into 2 groups, fasted for 24 h or refed for 5 h after a 24 h fasting. The mRNA concentrations for SREBPs and other lipogenic genes in the liver were determined by quantitative real time PCR. The hepatic mRNA relative abundance of lipogenic genes and genes involved in cholesterol synthesis were significantly greater (p<0.001) in the refed broilers. Similar results were demonstrated with Northern analysis. The data suggest that in the liver of fasted broilers, genes associated with lipogenesis and cholesterol biosynthesis were inhibited. Indeed, the mRNA concentrations for fatty acid synthase (FAS), malic enzyme, and stearoyl coenzyme A desaturase were almost undetectable after the 24 h fasting. The data also demonstrated that the expression of lipogenic genes coordinate well as a group during the refeeding period. Second, three small interfering RNA (siRNA) oligonucleotides against SREBP1 were designed to be used in transfecting a chicken hepatocarcinoma cell line LMH. One of the three siRNAs effectively reduced SREBP1 mRNA concentration (p<0.01). The acetyl coenzyme A carboxylase(alpha) (ACC(alpha)) mRNA was also significantly reduced by the SREBP1 siRNA treatment, suggesting that SREBP1 can upregulate the expression of this lipogenic gene. This siRNA, however, did not affect the mRNA for FAS. Taken together, the RNA interference study showed that SREBP1 has the ability to regulate the expression of ACC(alpha). This study has helped us understand more about the function of SREBP1 and the physiology of the broiler chickens.

Serum amyloid A protein regulates the expression of porcine genes related to lipid metabolism.

Serum amyloid A protein (SAA) is an apolipoprotein that can replace apolipoprotein A1 (apoA1) as the major apolipoprotein of HDL. Porcine hepatic SAA mRNA is increased by dietary docosahexaenoic acid (DHA) treatment. The purpose of this study was to investigate the role of SAA protein in regulating gene expression related to lipid metabolism in pigs. First, we demonstrated that the 100-micromol/L DHA treatment increased SAA and apoA1 mRNA expression in porcine hepatic cell cultures (P < 0.05). Secondly, we produced porcine SAA recombinant protein and found that the addition of SAA to porcine preadipocytes in culture stimulated interleukin-6 (IL-6) mRNA expression (P < 0.05), indicating a similar biological function of porcine SAA and human SAA. We also found PPARalpha and PPARgamma mRNA were decreased (40 and 60%, respectively) in differentiated adipocytes after treatment with 2 mumol/L SAA. SAA treatment also increased inflammatory cytokine gene expression (IL-6 and tumor necrosis factor alpha) and glycerol release (P < 0.05), indicating increased lipolysis. Because the expression of perilipin, a lipid droplet-protective protein, was reduced by the SAA treatment, we hypothesized that SAA increased lipolysis by decreasing the expression of perilipin, which would then allow an increase in hormone sensitive lipase activity. In conclusion, we demonstrated that the DHA-induced SAA gene expression decreased PPAR expression and consequently downregulated the expression of several genes involved in lipid metabolism. Accordingly, SAA may play a critical role in mediating the function of dietary DHA on lipid metabolism and could be a factor in regulating obesity.