Evidence-based guideline of the German Nutrition Society: fat intake and prevention of selected nutrition-related diseases

As nutrition-related chronic diseases have become more and more frequent, the importance of dietary prevention has also increased. Dietary fat plays a major role in human nutrition, and modification of fat and/or fatty acid intake could have a preventive potential. The aim of the guideline of the German Nutrition Society (DGE) was to systematically evaluate the evidence for the prevention of the widespread diseases obesity, type 2 diabetes mellitus, dyslipoproteinaemia, hypertension, metabolic syndrome, coronary heart disease (CHD), stroke, and cancer through the intake of fat or fatty acids. The main results can be summarized as follows: it was concluded with convincing evidence that a reduced intake of total and saturated fat as well as a larger intake of polyunsaturated fatty acids (PUFA) at the expense of saturated fatty acids (SFA) reduces the concentration of total and low-density lipoprotein cholesterol in plasma. Furthermore, there is convincing evidence that a high intake of trans fatty acids increases risk of dyslipoproteinaemia and that a high intake of long-chain polyunsaturated n-3 fatty acids reduces the triglyceride concentration in plasma. A high fat intake increases the risk of obesity with probable evidence when total energy intake is not controlled for (ad libitum diet). When energy intake is controlled for, there is probable evidence for no association between fat intake and risk of obesity. A larger intake of PUFA at the expense of SFA reduces risk of CHD with probable evidence. Furthermore, there is probable evidence that a high intake of long-chain polyunsaturated n-3 fatty acids reduces risk of hypertension and CHD. With probable evidence, a high trans fatty acid intake increases risk of CHD. The practical consequences for current dietary recommendations are described at the end of this article

Analysis of Human TAAR8 and Murine Taar8b Mediated Signaling Pathways and Expression Profile

The thyroid hormone derivative 3-iodothyronamine (3-T1AM) exerts metabolic effects in vivo that contradict known effects of thyroid hormones. 3-T1AM acts as a trace amine-associated receptor 1 (TAAR1) agonist and activates Gs signaling in vitro. Interestingly, 3-T1AM-meditated in vivo effects persist in Taar1 knockout-mice indicating that further targets of 3-T1AM might exist. Here, we investigated another member of the TAAR family, the only scarcely studied mouse and human trace-amine-associated receptor 8 (Taar8b, TAAR8). By RT-qPCR and locked-nucleic-acid (LNA) in situ hybridization, Taar8b expression in different mouse tissues was analyzed. Functionally, we characterized TAAR8 and Taar8b with regard to cell surface expression and signaling via different G-protein-mediated pathways. Cell surface expression was verified by ELISA, and cAMP accumulation was quantified by AlphaScreen for detection of Gs and/or Gi/o signaling. Activation of G-proteins Gq/11 and G12/13 was analyzed by reporter gene assays. Expression analyses revealed at most marginal Taar8b expression and no gender differences for almost all analyzed tissues. In heart, LNA-in situ hybridization demonstrated the absence of Taar8b expression. We could not identify 3-T1AM as a ligand for TAAR8 and Taar8b, but both receptors were characterized by a basal Gi/o signaling activity, a so far unknown signaling pathway for TAARs

Inverse agonistic action of 3-iodothyronamine at the human trace amine-associated receptor 5.

Application of 3-iodothyronamine (3-T1AM) results in decreased body temperature and body weight in rodents. The trace amine-associated receptor (TAAR) 1, a family A G protein-coupled receptor, is a target of 3-T1AM. However, 3-T1AM effects still persist in mTaar1 knockout mice, which suggest so far unknown further receptor targets that are of physiological relevance. TAAR5 is a highly conserved TAAR subtype among mammals and we here tested TAAR5 as a potential 3-T1AM target. First, we investigated mouse Taar5 (mTaar5) expression in several brain regions of the mouse in comparison to mTaar1. Secondly, to unravel the full spectrum of signaling capacities, we examined the distinct Gs-, Gi/o-, G12/13-, Gq/11- and MAP kinase-mediated signaling pathways of mouse and human TAAR5 under ligand-independent conditions and after application of 3-T1AM. We found overlapping localization of mTaar1 and mTaar5 in the amygdala and ventromedial hypothalamus of the mouse brain. Second, the murine and human TAAR5 (hTAAR5) display significant basal activity in the Gq/11 pathway but show differences in the basal activity in Gs and MAP kinase signaling. In contrast to mTaar5, 3-T1AM application at hTAAR5 resulted in significant reduction in basal IP3 formation and MAP kinase signaling. In conclusion, our data suggest that the human TAAR5 is a target for 3-T1AM, exhibiting inhibitory effects on IP3 formation and MAP kinase signaling pathways, but does not mediate Gs signaling effects as observed for TAAR1. This study also indicates differences between TAAR5 orthologs with respect to their signaling profile. In consequence, 3-T1AM-mediated effects may differ between rodents and humans

Investigation of Naturally Occurring Single-Nucleotide Variants in Human TAAR1

Activation of trace amine-associated receptor 1 (TAAR1) in endocrine pancreas is involved in weight regulation and glucose homeostasis. The purpose of this study was the identification and characterization of potential TAAR1 variants in patients with overweight/obesity and disturbed glucose homeostasis. Screening for TAAR1 variants was performed in 314 obese or overweight patients with impaired insulin secretion. The detected variants were functionally characterized concerning TAAR1 cell surface expression and signaling properties and their allele frequencies were determined in the population-based Study of Health in Pomerania (SHIP). Three heterozygous carriers of the single nucleotide missense variants p.Arg23Cys (R23C, rs8192618), p.Ser49Leu (S49L, rs140960896), and p.Ille171Leu (I171L, rs200795344) were detected in the patient cohort. While p.Ser49Leu and p.Ille171Leu were found in obese/overweight patients with slightly impaired glucose homeostasis, p.Arg23Cys was identified in a patient with a complete loss of insulin production. Functional in vitro characterization revealed a like wild-type function for I171L, partial loss of function for S49L and a complete loss of function for R23C. The frequency of the R23C variant in 2018 non-diabetic control individuals aged 60 years and older in the general population-based SHIP cohort was lower than in the analyzed patient sample. Both variants are rare in the general population indicating a recent origin in the general gene pool and/or the consequence of pronounced purifying selection, in line with the obvious detrimental effect of the mutations. In conclusion, our study provides hints for the existence of naturally occurring TAAR1 variants with potential relevance for weight regulation and glucose homeostasis

MAP kinase activation and G_s signaling parameters of wild type TAAR5 and chimeric receptors.

<p><b>(A)</b> HEK293 cells expressing mouse or human TAAR5 or chimeric receptors (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0117774#pone.0117774.t002" target="_blank">Table 2</a> for details) were stimulated with 100 μM DMEA. The cAMP accumulation was measured by competitive cAMP assay based on AlphaScreen technology. Results are depicted as either fold over basal mock or fold over DMEA stimulated mock transfection. Data are shown as mean ± SEM from n ≥ 3 independent experiments with three or more replicates. Statistical analyses were carried out with an unpaired two-tailed Welch-corrected t-test; ***p ≤ 0.001, compared to the respective basal activity. <b>(B)</b> MAP kinase activation was reported by luciferase activity in a luciferase reporter gene assay (SRE-luc). HEK293 cells were co-transfected with a reporter construct containing a serum response element and the firefly luciferase reporter gene, and the different receptor constructs. Cells were stimulated with 10 μM 3-T₁AM and SRE-luc levels were determined. Results are presented as mean ± SEM as either fold over basal mock transfection for basal value or fold over 3-T₁AM-stimulated mock. An unpaired two-tailed Welsh-corrected t-test was performed for statistical analyses; *p ≤ 0.05.</p

Signaling parameters of human and murine TAAR5 after treatment with 3-T₁AM.

<p><b>(A)</b> HEK293 cells expressing human TAAR1 were stimulated with 10 μM 3-T₁AM. For G_s signal determination cAMP accumulation was measured. Results are depicted as fold over basal mock or fold over 3-T₁AM-stimulated mock. Data is shown as mean ± SEM from n ≥ 3 independent experiments with 3 or more replicates. 3-T₁AM is a potent agonist for hTAAR1 (**p < 0.01). Statistical analysis was carried out with an unpaired two-tailed Welch-corrected t-test. <b>(B)</b> HEK293 cells transiently expressing hTAAR5 or mTaar5 were stimulated with 10 μM 3-T₁AM and IP3-luc levels were determined. Results are presented as either fold over basal mock transfection for basal value or fold over 3-T₁AM stimulated mock. An unpaired two-tailed Welsh-corrected t-test was used for statistical analyses, **p ≤ 0.01. Data are obtained from 3 to 6 independent experiments measured in at least triplicates and are shown as mean ± SEM. <b>(C)</b> Human TAAR5 was stimulated with 3-T₁AM concentrations ranging from 1 nM to 100 μM. The concentration-dependent IP3-luc signaling curve indicated the inverse agonism of 3-T₁AM at hTAAR5 with an EC₅₀ value of 4.4 ± 0.9 μM. <b>(D)</b> MAP kinase activation was determined by luciferase activity in a luciferase reporter gene assay (SRE-luc). HEK293 cells were co-transfected with a reporter construct containing a serum response element linked to the firefly luciferase reporter gene and in combination with the different receptor constructs, respectively. Cells were stimulated with 10 μM 3-T₁AM and SRE-luc levels were determined. Results are presented as mean ± SEM as either fold over basal mock transfection for basal value or fold over 3-T₁AM-stimulated mock. An unpaired two-tailed Welsh-corrected t-test was performed for statistical analyses; *p ≤ 0.05. <b>(E)</b> Cell surface expression studies of hTAAR5 were conducted in COS-7 cells for 6 hours after stimulation with or without 10 μM 3-T₁AM using an ELISA. Results are depicted as mean ± SEM obtained from 3 independent assays measured in 4 replicates. Data are presented as fold over basal hTAAR5. An unpaired two-tailed t-test with Welch-correction was performed.</p