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Dietary fatty acids play important roles in the regulation of fat accumulation or metabolic phenotype of adipocytes, either as brown or beige fat. However, a systematic comparison of effects of diets with different composition of 18-C fatty acids on browning/beiging phenotype has not been done. In this study, we compared the effects of different dietary fats, rich in specific 18-carbon fatty acids, on thermogenesis and lipid metabolism. Male C57BL/6 mice were fed a control diet containing 5.6% kcal fat from lard and 4.4% kcal fat from soybean oil (CON) or high-fat diets (HFD) containing 25% kcal from lard and 20% kcal fat from shea butter (stearic acid-rich fat; SHB), olive oil (oleic acid-rich oil; OO), safflower oil (linoleic acid-rich oil; SFO), or soybean oil (mixed oleic, linoleic, and α-linolenic acids; SBO) ad libitum for 12 weeks, with or without a terminal 4-h norepinephrine (NE) treatment. When compared to SHB, feeding OO, SFO, and SBO resulted in lower body weight gain. The OO fed group had the highest thermogenesis level, which resulted in lower body fat accumulation and improved glucose and lipid metabolism. Feeding SFO downregulated expression of lipid oxidation-related genes and upregulated expression of lipogenic genes, perhaps due to its high n-6:n-3 ratio. In general, HFD-feeding downregulated Ucp1 expression in both subcutaneous and epididymal white adipose tissue, and suppressed NE-induced Pgc1a expression in brown adipose tissue. These results suggest that the position of double bonds in dietary fatty acids, as well as the quantity of dietary fat, may have a significant effect on the regulation of oxidative and thermogenic conditions in vivo.

The objective of the current study was to determine if garlic-derived diallyl disulfide (DADS) and diallyl trisulfide (DATS) could mitigate oxidative and endotoxin stress, using an intestinal porcine epithelial cell (IPEC-J2) model. The experiment was arranged as a 2 × 2 × 2 factorial of DADS + DATS (0 or 18 µM), pro-oxidant stressor (hydrogen peroxide at 0 or 100 µM), and endotoxin stressor (lipopolysaccharide [LPS] at 0 or 10 µg/mL) with 8 replicates per treatment. Cells were incubated with DADS + DATS for 18 h, LPS for 6 h, then with hydrogen peroxide for 3 h. Gene expression was measured by RT-PCR for cytokines, interleukin 8 (IL-8) and tumor necrosis factor α (TNF-α), and tight junction proteins, claudin 1 (CL-1), occludin (OC), and zonula occludens 1 (ZO-1). Trans-epithelial electrical resistance (TEER), the antioxidant enzyme catalase, and apical secretion of IL-8 protein into the incubation medium was also measured. There was an increase ( < 0.01) in TNF-α and IL-8 gene expression due to LPS, although there was no effect of hydrogen peroxide or DADS + DATS. Furthermore, there was a tendency for an increase ( = 0.08) in ZO-1 gene expression due to DADS + DATS. Treatment with DADS + DATS and hydrogen peroxide did not affect TEER, although there was a decrease ( = 0.02) in TEER with LPS incubation. Treatment of cells with hydrogen peroxide reduced catalase activity ( < 0.01), which was restored with pre-incubation of DADS + DATS ( < 0.10). There was an increase ( < 0.01) in IL-8 secretion due to LPS, which was further augmented ( < 0.01) by pre-incubation with DADS + DATS. Based on the results from the current study, DADS + DATS can ameliorate oxidative effects of hydrogen peroxide, as well as alter IL-8 secretion in LPS-treated IPEC-J2 cells.

Short chain fatty acids (SCFA), products of microbial fermentation of dietary fiber, exert multiple metabolic effects in cells. Previously, we had demonstrated that soluble fiber influenced fat mass accumulation, gut microbial community structure and SCFA production in pigs. The current study was designed to identify effects of SCFA treatment during adipogenic differentiation of porcine stromovascular cells on lipid metabolism and adipokine expression. Differentiating cells were treated with varying concentrations of butyrate. Results show that butyrate treatment enhanced adipogenesis and lipid accumulation, perhaps through upregulation of glucose uptake and de novo lipogenesis and other mechanisms that include induction of SREBP-1c, C/EBPα/β, GLUT4, LPL, PPARγ, GPAT4, DGAT1 and DGAT2 expression. In addition, butyrate induced adiponectin expression, resulting in activation of downstream target genes, such as AMPK and AKT. Activation of AMPK by butyrate led to phosphorylation of ACC. Although increased ACO gene expression was seen with butyrate treatment, experiments with the peroxisomal fatty acid inhibitor, thioridazine, suggest that butyrate may have an inhibitory effect on peroxisomal fatty acid oxidation. Our studies also provide evidence that butyrate may inhibit lipolysis, perhaps in an FFAR3-dependent manner. Therefore, this study presents a novel paradigm for butyrate action in adipocytes and shows that adipocytes are capable of utilizing butyrate, leading to increased expression of adiponectin for enhanced glucose uptake and improved insulin sensitivity.

Obesity results in reduced differentiation potential of adipocytes leading to adipose tissue insulin resistance. Elevated proinflammatory cytokines from adipose tissue in obesity, such as TNFα have been implicated in the reduced adipocyte differentiation. Other mediators of reduced adipocyte differentiation include TGFβ and wnt proteins. Although some overlap exists in the signaling cascades of the wnt and TGFβ pathways it is unknown if TGFβ or wnt proteins reciprocally induce the expression of each other to maximize their biological effects in adipocytes. Therefore, we investigated the possible involvement of TGFβ signaling in wnt induced gene expression and vice versa in 3T3-L1 adipocyte. Effect of TGFβ and Wnt pathways on differentiation was studied in preadipocytes induced to differentiate in the presence of Wnt3a or TGFβ1 and their inhibitors (FZ8-CRD and SB431542, respectively). Regulation of intracellular signaling and gene expression was also studied in mature adipocytes. Our results show that both TGFβ1 and Wnt3a lead to increased accumulation of β-catenin, phosphorylation of AKT and p44/42 MAPK. However, differences were found in the pattern of gene expression induced by the two proteins suggesting that distinct, but complex, signaling pathways are activated by TGFβ and wnt proteins to independently regulate adipocyte function.

Weaned pigs often experience growth reduction immediately after weaning due to multiple stress factors associated with weaning. We tested the effect of prenatal and postnatal butyrate supplementation on growth performance of piglets. In study 1, piglets were orally gavaged with 0.3% butyrate from day 4 after birth to weaning (day 21). Butyrate increased ADG by 13% compared to saline treated control. Expression of peroxisome proliferator activated receptor gamma coactivator 1 alpha (PGC-1α) was higher in muscle, adipose tissue, and ileum of butyrate-supplemented animals. Also, peroxisome proliferator activated receptor alpha (PPARα) was induced (P < 0.05) in the subcutaneous adipose tissue (SCAT) and muscle (longissimus dorsi [LD]) of butyrate-supplemented piglets. In vitro, butyrate increased (P < 0.05) fatty acid oxidation in primary adipocytes and suppressed basal lipolysis by 62% compared to untreated cells. Butyrate suppressed (P < 0.05) lipogenesis ((14)C-glucose incorporation into lipids) in adipocytes. This was accompanied by an approximately 30% reduction in the mRNA expression of fatty acid synthase (P < 0.05) in butyrate-treated cells vs. controls. Piglets born to sows that were supplemented with 0.3% butyrate during the last trimester of gestation had a 15% higher (P < 0.05) body weight at 12 wk than controls. In summary, butyrate supplementation to gestating sows and piglets enhanced postweaning growth performance, which may be mediated by increased substrate oxidation in butyrate treated animals.

Obesity promotes increased secretion of a number of inflammatory factors from adipose tissue. These factors include cytokines and very lately, extracellular matrix components (ECM). Biglycan, a small leucine rich proteoglycan ECM protein, is up-regulated in obesity and has recently been recognized as a pro-inflammatory molecule. However, it is unknown whether biglycan contributes to adipose tissue dysfunction. In the present study, we characterized biglycan expression in various adipose depots in wild-type mice fed a low fat diet (LFD) or obesity-inducing high fat diet (HFD). High fat feeding induced biglycan mRNA expression in multiple adipose depots. Adiponectin is an adipokine with anti-inflammatory and insulin sensitizing effects. Due to the importance of adiponectin, we examined the effect of biglycan on adiponectin expression. Comparison of adiponectin expression in biglycan knockout (bgn(-/0)) and wild-type (bgn(+/0)) reveals higher adiponectin mRNA and protein in epididymal white adipose tissue in bgn(-/0) mice, as well higher serum concentration of adiponectin, and lower serum insulin concentration. On the contrary, knockdown of biglycan in 3T3-L1 adipocytes led to decreased expression and secretion of adiponectin. Furthermore, treatment of 3T3-L1 adipocytes with conditioned medium from biglycan treated macrophages resulted in an increase in adiponectin mRNA expression. These data suggest a link between biglycan and adiponectin expression. However, the difference in the pattern of regulation between in vivo and in vitro settings reveals the complexity of this relationship.

Two in vivo studies were conducted with pigs to determine the effects of exogenous leptin on the expression of peroxisome proliferator activated receptors (PPAR), and on serum concentrations of selected metabolites and hormones. Initially, leptin was administered i.m. to young pigs for 15 days at 0 (control), 0.003 (low), 0.01 (medium) and 0.03 (high) mg. kg(-1). day(-1). There was no leptin effect on serum glucose (P > 0.84), triglycerides (P > 0.69), non-esterified fatty acids (NEFA, P > 0.53), or glycerol (P > 0.33). Leptin at the intermediate and high doses depressed adipose expression of both PPARgamma1 (P < 0.06) and PPARgamma2 (P < 0.01). In a second study, we used a paired-feeding experimental design to determine the effects of a higher dose of leptin (0.05 mg. kg(-1). day(-1)) on serum metabolites and PPAR expression in selected tissues. At this dose, leptin increased (P < 0.0001) serum NEFA concentrations relative to both the ad libitum and pair-fed control groups. However, in this study, there was no difference in the expression of PPARgamma1 in adipose tissue, but PPARgamma2 mRNA was upregulated by leptin (P < 0.08). In contrast, leptin had no impact on the expression of PPARalpha in liver, skeletal muscle or adipose tissue. Adipose tissue explants were also incubated with leptin to assess the effect on PPARgamma expression, in vitro. The abundance of PPARgamma1 mRNA (P < 0.05) was increased after 24 hr of exposure, but the effect of leptin on gamma2 was not significant (P > 0.24). The lipolytic effect of leptin was also evaluated in vitro using isolated adipocytes. In keeping with the increase in serum NEFA concentrations in vivo, leptin stimulated lipolysis in vitro, increasing glycerol concentrations in the medium to about 219% of that in basal (non-treated) culture medium after 8 hr of incubation. Collectively, the data presented herein indicate that leptin modulates lipid metabolism in the pig, but that PPARalpha expression is not a parallel target of leptin as it is in rodent models. The regulation of PPARgamma by leptin seems complex in that it varied in relation to dose in vivo, and may be impacted by in vitro vs. in vivo circumstances.

A combination of in vivo and in vitro experiments were performed to determine the extent to which exogenous leptin regulates serum growth hormone (GH) and insulin-like growth factor I (IGF-1) concentrations, and the abundance of IGF-1 mRNA in major peripheral tissues. Initially (Experiment 1), a recombinant human leptin analog was administered i.m. to young growing pigs (approximately 27 kg body weight) for 15 days at 0 (control), 0.003, 0.01 and 0.03 mg. kg(-1). day(-1). Although there was no sustained effect of leptin on serum GH, there was a reduction (P < 0.02) in serum IGF-1 at the intermediate dose that paralleled a decrease (P < 0.09) in hepatic IGF-1 expression. Leptin, at these doses, did not reduce feed intake (P > 0.57), nor was there an effect of leptin on dietary nitrogen retention (P > 0.97). In a second experiment, pigs were injected with vehicle or a higher dose of leptin (0.05 mg. kg(-1). day(-1)) for 14 days. A third treatment group was injected with vehicle and pair-fed to the intake of the group treated with leptin. In this study, exogenous leptin resulted in a sustained increase in serum leptin (P < 0.0001) and reduction in feed intake of approximately 30% (P < 0.0001). Serum IGF-1 was depressed in both the leptin-treated and pair-fed groups, relative to the group allowed ad-libitum intake (P < 0.01). Furthermore, there was no difference among treatments in the relative abundance of IGF-1 mRNA in skeletal muscle (P > 0.42) or adipose tissue (P > 0.26), and liver mRNA abundance was actually increased (P < 0.01) by leptin, despite the lower feed intake. Finally, to determine whether leptin altered the secretion of IGF-1 by isolated pig hepatocytes, primary cultures were incubated with leptin for 24 to 48 hr (Experiment 3). Leptin (100 nM) caused a sharp reduction (P < 0.0001) in dexamethasone-induced IGF-1 secretion at 24 hr (47% reduction) and at 48 hr (40% reduction). Collectively, these data indicate that leptin may regulate hepatic IGF-1 production in the pig, independent of GH, but that hepatocyte sensitivity to leptin may be depend on dose and in vitro vs. in vivo conditions.