Time course of adiponectin level in the explants incubation medium and serum.

<p>At the age of 5w, 6w and 7w, adiponectin level in the explants incubation medium (EIM) of the db/db mice was significantly lower than that of lean mice(P<0.01) (Fig. 8A), indicating that adiponectin production from the epididymal adipose tissue was decreased in db/db mice; Adiponectin in the serum was comparable between db/db mice and lean mice at each point (Fig. 8B).</p

Treatment with Rosiglitazone increased the serum resistin level of db/db mice.

<p>Db/db mice were treated with PBS (control) or Rosiglitazone (20 mg/kg/day) by daily intra-gastric gavage for 4 weeks. The db/db mice treated with Rosiglitazone (Rosi) had higher level of serum resistin <i>vs</i> control(*P<0.05).</p

Effects of Rosiglitazone on the protein production and gene expression of resistin in adipocytes differentiated from 3T3-L1.

<p>Fully differentiated 3T3-L1 adipocytes were cultured with or without Rosiglitazone (10 uM) for 48 h. The resistin in the conditioned medium was measured (Fig. 7A). Total RNA purified from these samples was subjected to quantitative PCR analysis to determine the mRNA levels of resistin. Data were normalized against 18 s ribosomal RNA. For comparison, the levels of the resistin without Rosiglitazone were arbitrarily set at 1(Fig. 7B). The results were repeated for three times and expressed as mean ±S. <i>vs</i> control, *p<0.05**p<0.01.</p

The resistin gene expression in the epididymal adipose tissue at the age of 5 weeks.

<p>Total RNA purified from these samples was subjected to quantitative PCR analysis to determine the mRNA levels of resistin. Data were normalized against 18s ribosomal RNA. For comparison, the gene expression levels in the lean control mice were arbitrarily set at 1. **p<0.01 <i>vs</i> lean control.</p

The time course of body weight (BW) and total adipose tissue (tAT) percentage of BW.

<p>At the age of 5 week, BW was comparable between db/db mice and lean control. Db/db mice older than 6W had higher BW than lean control (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065543#pone-0065543-g001" target="_blank">Figure 1A</a>). At each time point, tAT(all AT from epididymal, perirenal and abodomal) percentage of body weight(BW) from db/db mice were significantly higher than lean control(<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0065543#pone-0065543-g001" target="_blank">Figure 1B</a>), indicating that fat tissue mass increased markedly in db/db mice from 5 weeks old.*P<0.01,compared with lean control at the same age.</p

Time course of resistin level in the explants incubation medium (EIM) and its relation to serum resistin level.

<p>At each time point, resistin production was decreased in db/db mice(<i>vs</i> lean control at the same age *P<0.05, **P<0.01, Fig. 4A); Fig. 4B showed the positive correlation between the resistin level in the explants incubation medium and the resistin level in the serum (r = 0.844 P = 0.000).</p

The Critical Role of Astragalus Polysaccharides for the Improvement of PPRAα-Mediated Lipotoxicity in Diabetic Cardiomyopathy

<div><h3>Background</h3><p>Obesity-related diabetes mellitus leads to increased myocardial uptake and oxidation of fatty acids, resulting in a form of cardiac dysfunction referred to as lipotoxic cardiomyopathy. We have shown previously that Astragalus polysaccharides (APS) administration was sufficient to improve the systemic metabolic disorder and cardiac dysfunction in diabetic models.</p> <h3>Methodology/Principal Findings</h3><p>To investigate the precise role of APS therapy in the pathogenesis of myocardial lipotoxity in diabetes, db/db diabetic mice and myosin heavy chain (MHC)- peroxisome proliferator-activated receptor (PPAR) α mice were characterized and administrated with or without APS with C57 wide- type mice as normal control. APS treatment strikingly improved the myocyte triacylglyceride accumulation and cardiac dysfunction in both db/db mice and MHC-PPARα mice, with the normalization of energy metabolic derangements in both db/db diabetic hearts and MHC-PPARα hearts. Consistently, the activation of PPARα target genes involved in myocardial fatty acid uptake and oxidation in both db/db diabetic hearts and MHC-PPARα hearts was reciprocally repressed by APS administration, while PPARα-mediated suppression of genes involved in glucose utilization of both diabetic hearts and MHC-PPARα hearts was reversed by treatment with APS.</p> <h3>Conclusions</h3><p>We conclude that APS therapy could prevent the development of diabetic cardiomyopathy through a mechanism mainly dependent on the cardiac PPARα-mediated regulatory pathways.</p> </div

Gene expression of PPARα target genes in hearts of mice at different age.

<p>Cardiac tissues from db/db mice and MHC-PPARα mice with/without APS administration were detected for the gene expression at the age of 5 weeks (A) and 20 weeks (B-H) respectively, with C57 mice as normal control. Myocardial mRNA levels of PPARα target genes were determined by real-time RT-PCR analysis, encoding PPARα (B), M-CPF 1(C), ACO (D), FATP 1 (E), FACS 1(F), GLUT4 (G) and PDK4 (H). The relative gene expression of db/db mice and MHC-PPARα mice was normalized to the value of C57 wide-type mice, and expressed as fold change. All groups n = 6. Bars represent means±S.E.M. *<i>P</i><0.05 vs. C57 mice, # <i>P</i><0.05 vs. untreated db/db mice, † <i>P</i><0.05 vs. untreated MHC-PPARα mice. M-CPF1, muscle-typecarnitine palmitoyltransferase 1; ACO, acyl-CoA oxidase; FATP 1, fatty acid trasport protein-1; FACS 1, fatty acyl-CoA synthetase 1; GLUT4, glucose transporter 4; PDK4, pyruvate dehydrogenase kinase 4.</p

Intramyocardial lipid accumulation in mice.

<p>Myocardial triacylglyceride levels (A) in db/db mice and MHC-PPARα mice with or without APS administration at 20 week-old were determined by ESI/MS. Mean levels of TAG-associated fatty acid with chain length of 16∶0 (B), 18∶0 (C), 18∶1 (D), and 18∶2 (E) in the hearts of each group were shown respectively. (F) representative images of myocardium stained with oil red O. All groups n = 8. Bars represent means±S.E.M. *<i>P</i><0.05 vs. C57 mice, # <i>P</i><0.05 vs. untreated db/db mice, † <i>P</i><0.05 vs. untreated MHC-PPARα mice.</p

Serum biometric parameters of mice at different age.

<p>Serum glucose, insulin, nonesterified fatty acid (FFA) and triacylglyceride (TAG) were detected in db/db mice and MHC-PPARα mice with or without APS administration at the age of 5 weeks (A) and 20 weeks (B–E) respectively, with C57 wide-type mice as normal control. All groups n = 8. Bars represent means±S.E.M. *<i>P</i><0.05 vs. C57 mice, # <i>P</i><0.05 vs. untreated db/db mice, † <i>P</i><0.05 vs. untreated MHC-PPARα mice.</p