We sought to determine the role of adipocyte death in obesity-induced adipose tissue (AT) inflammation and obesity complications.
Male C57BL/6 mice were fed a high-fat diet for 20 weeks to induce ...obesity. Every 4 weeks, insulin resistance was assessed by intraperitoneal insulin tolerance tests, and epididymal (eAT) and inguinal subcutaneous AT (iAT) and livers were harvested for histological, immunohistochemical, and gene expression analyses.
Frequency of adipocyte death in eAT increased from <0.1% at baseline to 16% at week 12, coincident with increases in 1) depot weight; 2) AT macrophages (ATM Phi s) expressing F4/80 and CD11c; 3) mRNA for tumor necrosis factor (TNF)-alpha, monocyte chemotactic protein (MCP)-1, and interleukin (IL)-10; and 4) insulin resistance. ATM Phi s in crown-like structures surrounding dead adipocytes expressed TNF-alpha and IL-6 proteins. Adipocyte number began to decline at week 12. At week 16, adipocyte death reached approximately 80%, coincident with maximal expression of CD11c and inflammatory genes, loss (40%) of eAT mass, widespread collagen deposition, and accelerated hepatic macrosteatosis. By week 20, adipocyte number was restored with small adipocytes, coincident with reduced adipocyte death (fourfold), CD11c and MCP-1 gene expression (twofold), and insulin resistance (35%). eAT weight did not increase at week 20 and was inversely correlated with liver weight after week 12 (r = -0. 85, P < 0.001). In iAT, adipocyte death was first detected at week 12 and remained <or=3%.
These results implicate depot-selective adipocyte death and M Phi-mediated AT remodeling in inflammatory and metabolic complications of murine obesity.
•High fat feeding increased phosphatidylinositol-3 kinase signaling.•Obese mice had altered xenobiotic metabolism gene expression.•Obesity induced altered microRNA expression.
Insulin regulates ...ovarian phosphatidylinositol-3-kinase (PI3 K) signaling, important for primordial follicle viability and growth activation. This study investigated diet-induced obesity impacts on: (1) insulin receptor (Insr) and insulin receptor substrate 1 (Irs1); (2) PI3K components (Kit ligand (Kitlg), kit (c-Kit), protein kinase B alpha (Akt1) and forkhead transcription factor subfamily 3 (Foxo3a)); (3) xenobiotic biotransformation (microsomal epoxide hydrolase (Ephx1), Cytochrome P450 isoform 2E1 (Cyp2e1), Glutathione S-transferase (Gst) isoforms mu (Gstm) and pi (Gstp)) and (4) microRNA's 184, 205, 103 and 21 gene expression. INSR, GSTM and GSTP protein levels were also measured. Obese mouse ovaries had decreased Irs1, Foxo3a, Cyp2e1, MiR-103, and MiR-21 but increased Kitlg, Akt1, and miR-184 levels relative to lean littermates. These results support that diet-induced obesity potentially impairs ovarian function through aberrant gene expression.
Under certain dietary situations, rumen biohydrogenation results in the production of unique fatty acids that inhibit milk fat synthesis. The first of these to be identified was trans-10, cis-12 ...conjugated linoleic acid (CLA), but others are postulated to contribute to diet-induced milk fat depression (MFD). Our objective was to examine the potential role of trans-9, cis-11 CLA in the regulation of milk fat. In a preliminary study, we used gas-liquid and high-performance liquid chromatography techniques to examine milk fat samples from a diet-induced MFD study and found that an increase in trans-9, cis-11 CLA corresponded to the decrease in milk fat yield. We investigated this further using a CLA enrichment of 9, 11 isomers to examine the biological effect of trans-9, cis-11 CLA on milk fat synthesis. Four rumen-fistulated Holstein cows were randomly assigned in a 4 x 4 Latin square experiment involving 5-d treatment periods and abomasal infusion of 1) ethanol (control), 2) a 9, 11 CLA mix (containing 32% trans-9, cis-11, 29% cis-9, trans-11, and 17% trans-9, trans-11), 3) a trans-9, trans-11 CLA supplement, and 4) a trans-10, cis-12 CLA supplement (positive control). The trans-9, trans-11 CLA and trans-10, cis-12 CLA supplements were of high purity (>90%), and all supplements were infused at a rate to provide 5 g/d of the CLA isomer of interest. Milk yield and dry matter intake did not differ among treatments. Compared with the control treatment, milk fat yield was reduced by 15% for the 9, 11 CLA mixture and by 27% for the trans-10, cis-12 CLA treatment. We also found that trans-9, trans-11 CLA had no effect on milk fat yield, and previous research has shown that milk fat yield is unaltered when cows are infused with cis-9, trans-11 CLA. When all treatments were considered, results suggested that trans-9, cis-11 was the CLA isomer in the 9, 11 CLA mix responsible for the reduction in milk fat synthesis, although the magnitude was less than that observed for trans-10, cis-12 CLA. Interestingly, trans-9, trans-11 CLA altered the milk fat desaturase index, further demonstrating that alterations in desaturase can occur independently of effects on milk fat synthesis. Overall, our investigations identified that an increase in milk fat content of trans-9, cis-11 CLA was associated with diet-induced MFD and provided evidence of a role for this isomer in MFD based on the 15% reduction in milk fat yield with abomasal infusion of a CLA enrichment that supplied 5 g/d of trans-9, cis-11 CLA.
Perilipin A is the most abundant phosphoprotein on adipocyte lipid droplets and is essential for lipid storage and lipolysis. Perilipin null mice exhibit diminished adipose tissue, elevated basal ...lipolysis, reduced catecholamine-stimulated lipolysis, and increased insulin resistance. To understand the physiological consequences of increased perilipin expression in vivo, we generated transgenic mice that overexpressed either human or mouse perilipin using the adipocyte-specific aP2 promoter/enhancer. Phenotypes of female transgenic and wild-type mice were characterized on chow and high-fat diets (HFDs). When challenged with an HFD, transgenic mice exhibited lower body weight, fat mass, and adipocyte size than wild-type mice. Expression of oxidative genes was increased and lipogenic genes decreased in brown adipose tissue of transgenic mice. Basal and catecholamine-stimulated lipolysis was decreased and glucose tolerance significantly improved in transgenic mice fed a HFD. Perilipin overexpression in adipose tissue protects against HFD-induced adipocyte hypertrophy, obesity, and glucose intolerance. Alterations in brown adipose tissue metabolism may mediate the effects of perilipin overexpression on body fat, although the mechanisms by which perilipin overexpression alters brown adipose tissue metabolism remain to be determined. Our findings demonstrate a novel role for perilipin expression in adipose tissue metabolism and regulation of obesity and its metabolic complications.
Milk fat depression (MFD) is a naturally occurring condition in dairy cows where milk fat synthesis is inhibited by intermediates of ruminal biohydrogenation. One of these bioactive fatty acids (FA), ...trans-10, cis-12 conjugated linoleic acid (CLA), decreases milk fat synthesis through transcriptional downregulation of genes involved in mammary lipid synthesis. Energy partitioning during MFD is not well characterized because of the complexity of observing energy metabolism in ruminant animals. To investigate energy partitioning during MFD, adipose tissue biopsies were taken from 4 cows arranged in a switchback design. Treatments were control and 4-d abomasal infusion of trans-10, cis-12 CLA (7.5 g/d). CLA decreased milk fat yield by 38% and milk fat content by 34%, but yields of milk and other milk components were unchanged. In contrast to reported changes in mammary tissue, adipose tissue expression of lipid synthesis enzymes, including lipoprotein lipase, FA synthase, stearoyl-CoA desaturase, and FA binding protein 4, was increased. Expression of regulators of lipid synthesis, including sterol-response element binding protein 1, thyroid hormone responsive spot 14, and PPARγ, also increased in adipose tissue. Thus, a CLA dose resulting in near maximal inhibition of mammary lipid synthesis resulted in increased expression of lipid synthesis-related genes in adipose tissue. A meta-analysis of intake response during CLA infusion was conducted to extend the investigation of energy metabolism during MFD. Voluntary intake decreased (P < 0.001) by 1.5 kg/d during CLA-induced MFD in the 14 studies analyzed, but the reduction in intake only partially accounts for the energy spared from reduced milk fat synthesis. Results are consistent with energy spared from the reduction in milk fat synthesis being partitioned toward adipose tissue fat stores during short-term MFD.
Obesity-associated low-grade systemic inflammation resulting from increased adipose mass is strongly related to the development of insulin resistance and type 2 diabetes as well as other metabolic ...complications. Recent studies have demonstrated that the obese metabolic state can be improved by ablating certain inflammatory signaling pathways. Tumor progression locus 2 (TPL2), a kinase that integrates signals from Toll receptors, cytokine receptors, and inhibitor of κ-B kinase-β is an important regulator of inflammatory pathways. We used TPL2 knockout (KO) mice to investigate the role of TPL2 in mediating obesity-associated inflammation and insulin resistance.
Male TPL2KO and wild-type (WT) littermates were fed a low-fat diet or a high-fat diet to investigate the effect of TPL2 deletion on obesity, inflammation, and insulin sensitivity.
We demonstrate that TPL2 deletion does not alter body weight gain or adipose depot weight. However, hyperinsulinemic euglycemic clamp studies revealed improved insulin sensitivity with enhanced glucose uptake in skeletal muscle and increased suppression of hepatic glucose output in obese TPL2KO mice compared with obese WT mice. Consistent with an improved metabolic phenotype, immune cell infiltration and inflammation was attenuated in the adipose tissue of obese TPL2KO mice coincident with reduced hepatic inflammatory gene expression and lipid accumulation.
Our results provide the first in vivo demonstration that TPL2 ablation attenuates obesity-associated metabolic dysfunction. These data suggest TPL2 is a novel target for improving the metabolic state associated with obesity.
Exercise enhances insulin sensitivity; it also improves adipocyte metabolism and reduces adipose tissue inflammation through poorly defined mechanisms. Fibroblast growth factor 21 (FGF21) is a ...pleiotropic hormone-like protein whose insulin-sensitizing properties are predominantly mediated via receptor signaling in adipose tissue (AT). Recently, FGF21 has also been demonstrated to have anti-inflammatory properties. Meanwhile, an association between exercise and increased circulating FGF21 levels has been reported in some, but not all studies. Thus, the role that FGF21 plays in mediating the positive metabolic effects of exercise in AT are unclear. In this study, FGF21-knockout (KO) mice were used to directly assess the role of FGF21 in mediating the metabolic and anti-inflammatory effects of exercise on white AT (WAT) and brown AT (BAT). Male FGF21KO and wild-type mice were provided running wheels or remained sedentary for 8 weeks (
= 9-15/group) and compared for adiposity, insulin sensitivity (i.e., HOMA-IR, Adipo-IR) and AT inflammation and metabolic function (e.g., mitochondrial enzyme activity, subunit content). Adiposity and Adipo-IR were increased in FGF21KO mice and decreased by EX. The BAT of FGF21KO animals had reduced mitochondrial content and decreased relative mass, both normalized by EX. WAT and BAT inflammation was elevated in FGF21KO mice, reduced in both genotypes by EX. EX increased WAT
gene expression, citrate synthase activity, COX I content and total AMPK content in WT but not FGF21KO mice. Collectively, these findings reveal a previously unappreciated anti-inflammatory role for FGF21 in WAT and BAT, but do not support that FGF21 is necessary for EX-mediated anti-inflammatory effects.
The efficacy of conjugated linoleic acid (CLA) supplements containing trans-10, cis-12 for reducing milk fat synthesis has been well documented in dairy cows, but studies with other ruminant species ...are less convincing, and there have been no investigations of this in sheep. Therefore, the current study was designed to determine whether trans-10, cis-12 CLA would inhibit milk fat synthesis in sheep. Twenty multiparous ewes in early lactation were paired and randomly allocated to 2 treatments: grass hay plus concentrate either unsupplemented (control) or supplemented with lipid-encapsulated CLA to provide 2.4 g/d of trans-10, cis-12 CLA. The CLA dose was based on published responses of dairy cows extrapolated to ewes on a metabolic body weight basis. The experimental design was a 2-period crossover with 10-d treatment periods separated by a 10-d interval. Compared with the control, CLA supplementation reduced milk fat content from 6.4 to 4.9% and reduced fat yield from 95 to 80 g/d. The CLA treatment also increased milk yield from 1,471 to 1,611 g/d and increased protein yield from 68 to 73 g/d. Milk protein content and DMI were unaffected by treatment. The reduction in milk fat yield was due to decreases in both de novo fatty acid synthesis and uptake of preformed fatty acids. Milk fat content of trans-10, cis-12 CLA was < 0.01 and 0.12 g/100 g of fatty acids for the control and CLA treatments, respectively. The transfer efficiency of trans-10, cis-12 CLA from the dietary supplement into milk fat was 3.8%. Results of the present study demonstrate that a CLA supplement containing trans-10, cis-12 CLA reduces milk fat synthesis in lactating sheep in a manner similar to dairy cows when fed at an equivalent dose (metabolic body weight basis). Furthermore, the nutrients spared by the reduction in milk fat coincided with an increase in milk and milk protein yield.
In response to cold, norepinephrine (NE)-induced triacylglycerol hydrolysis (lipolysis) in adipocytes of brown adipose tissue (BAT) provides fatty acid substrates to mitochondria for heat generation ...(adaptive thermogenesis). NE-induced lipolysis is mediated by protein kinase A (PKA)-dependent phosphorylation of perilipin, a lipid droplet-associated protein that is the major regulator of lipolysis. We investigated the role of perilipin PKA phosphorylation in BAT NE-stimulated thermogenesis using a novel mouse model in which a mutant form of perilipin, lacking all six PKA phosphorylation sites, is expressed in adipocytes of perilipin knockout (Peri KO) mice. Here, we show that despite a normal mitochondrial respiratory capacity, NE-induced lipolysis is abrogated in the interscapular brown adipose tissue (IBAT) of these mice. This lipolytic constraint is accompanied by a dramatic blunting (∼70%) of the in vivo thermal response to NE. Thus, in the presence of perilipin, PKA-mediated perilipin phosphorylation is essential for NE-dependent lipolysis and full adaptive thermogenesis in BAT. In IBAT of Peri KO mice, increased basal lipolysis attributable to the absence of perilipin is sufficient to support a rapid NE-stimulated temperature increase (∼3.0°C) comparable to that in wild-type mice. This observation suggests that one or more NE-dependent mechanism downstream of perilipin phosphorylation is required to initiate and/or sustain the IBAT thermal response.
Short-term studies (< 5 d) involving abomasal infusion of a mixture of CLA isomers or pure trans-10, cis-12 CLA have demonstrated that supplements of conjugated linoleic acids (CLA) reduce milk fat ...synthesis during established lactation in dairy cows. Our objective was to assess longer term effects of supplementation during established lactation using a dietary supplement of rumen-protected CLA. Thirty Holstein cows were blocked by parity and received a dietary fat supplement of either Ca-salts of palm oil fatty acids (control) or a mixture of Ca-salts of palm oil fatty acids plus Ca-salts of CLA (CLA treatment). Supplements provided about 90 g/d of fatty acids and were topdressed on the TMR. The CLA supplement provided 30.4 g/d of CLA in which the predominant isomers were: trans-8, cis-10 (9.2%), cis-9, trans-11 (25.1%), trans-10, cis-12 (28.9%), and cis-11, trans-13 (16.1%). All cows were pregnant; treatments were initiated on d 79 of pregnancy (approximately 200 d prepartum) and continued for 140 d until dry off. Twenty-three cows completed the study; those receiving CLA supplement had a lower milk fat test (2.90 versus 3.80%) and a 23% reduction in milk fat yield (927 versus 1201 g/d). Intake of DM, milk yield, and the yield and content of true protein and lactose in milk were unaffected by treatment. Milk fat analysis indicated that the CLA supplement reduced the secretion of fatty acids of all chain lengths. However, effects were proportionally greater on short and medium chain fatty acids, thereby causing a shift in the milk fatty acid composition to a greater content of longer-chain fatty acids. Changes in body weight gain, body condition score, and net energy balance were not significant and imply no differences in cows fed the CLA supplement in replenishment of body reserves in late lactation. Likewise, maintenance of pregnancy, gestation length, and calf birth weight were unaffected by treatment. Overall, feeding a dietary supplement of rumen-protected CLA to pregnant cows over the last 140 d of the lactation cycle resulted in a marked reduction in milk fat content and yield, and a shift in milk fatty acid composition, but other milk components, DMI, maintenance of pregnancy, and cow well-being were unaffected.
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