Mouse PPAR-α(Peroxisome prolifeRator-activated receptor alpha) ELISA Kit

The peroxisome proliferator activated receptors (PPARs) are important drug targets in treatment of metabolic and inflammatory disorders. Fibrates, acting as PPAR alpha agonists, have been widely used lipid-lowering agents for decades. However, the currently available PPAR alpha targeting agents show low subtype-specificity and consequently a search for more potent agonists have emerged. In this study, previously isolated oxohexadecenoic acids from the marine algae Chaetoceros karianus were used to design a PPAR alpha-specific analogue. Herein we report the design, synthesis, molecular modelling studies and biological evaluations of the novel 3,5-disubstituted isoxazole analogue 6-(5-heptyl-1,2-oxazol-3-yl)hexanoic acid (1), named ADAM. ADAM shows a clear receptor preference and significant dose-dependent activation of PPAR alpha (EC50 = 47 mu M) through its ligand-binding domain (LBD). Moreover, ADAM induces expression of important PPAR gamma target genes, such as CPT1A, in the Huh7 cell line and primary mouse hepatocytes. In addition, ADAM exhibits a moderate ability to regulate PPAR alpha target genes and drive adipogenesis. Molecular modelling studies indicated that ADAM docks its carboxyl group into opposite ends of the PPAR alpha and -gamma LBD. ADAM interacts with the receptor-activating polar network of amino acids (Tyr501, His447 and Ser317) in PPAR alpha, but not in PPAR gamma LBD. This may explain the lack of PPAR gamma agonism, and argues for a PPARa-dependent adipogenic function. Such compounds are of interest towards developing new lipid-lowering remedies.

Objective:The fatty acid receptor 1 (FFAR1/GPR40) mediates fatty acid-dependent augmentation of glucose-induced insulin secretion (GIIS) in pancreatic beta-cells. Genetically engineered Ffar1-knockout/congenic mice univocally displayed impaired fatty acid-mediated insulin secretion, but in vivo experiments delivered controversial results regarding the function of FFAR1 in glucose homeostasis and liver steatosis. This study presents a new coisogenic mouse model carrying a point mutation in Ffar1 with functional consequence. These mice reflect the situations in humans in which point mutations can lead to protein malfunction and disease development. Methods: The Munich N-ethyl-N-nitrosourea (ENU) mutagenesis-derived F1 archive containing over 16,800 sperms and corresponding DNA samples was screened for mutations in the coding region of Ffar1. Two missense mutations (R258W and T146S) in the extracellular domain of the protein were chosen and homozygote mice were generated. The functional consequence of these mutations was examined in vitro in isolated islets and in vivo in chow diet and high fat diet fed mice. Results: Palmitate, 50 mu M, and the FFAR1 agonist TUG-469, 3 mu M, stimulated insulin secretion in islets of Ffar1(T146S/T146S) mutant mice and of wild-type littermates, while in islets of Ffar1(R258W/R258W) mutant mice, these stimulatory effects were abolished. Insulin content and mRNA levels of Ffar1, Glp1r, Ins2, Slc2a2, Ppara, and Ppard were not significantly different between wild-type and Ffar1(R258W/R258W) mouse islets. Palmitate exposure, 600 mu M, significantly increased Ppara mRNA levels in wild-type but not in Ffar1(R258W/R258W) mouse islets. On the contrary, Slc2a2 mRNA levels were significantly reduced in both wild-type and Ffar1(R258W/R258W) mouse islets after palmitate treatment. HFD feeding induced glucose intolerance in wild-type mice. Ffar1(R258W/R258W) mutant mice remained glucose tolerant although their body weight gain, liver steatosis, insulin resistance, and plasma insulin levels were not different from those of wild-type littermates. Worth mentioning, fasting plasma insulin levels were lower in Ffar1(R258W/R258W) mice. Conclusion: A point mutation in Ffar1 abrogates the stimulatory effect of palmitate on GIIS, an effect that does not necessarily translate to HFD-induced glucose intolerance.