Gene expression profile of peripheral blood mononuclear cells in mild to moderate obesity in dogs.

BACKGROUND: Molecular mechanisms and early diagnosis on the development of mild to moderate of canine obesity are not understood although recent dog obesity is a widespread problem. To understand the differences between normal weight and mild to moderate obesity, the purpose of this study is to investigate the gene expression profiles of peripheral blood mononuclear cells (PBMC) in dogs. METHODS: This study comprised a sample of 12 privately-owned Miniature Dachshund, which were divided into two groups (obese and control) based on body condition scores (BCS). Serum biochemical parameters and PBMC gene expression profiles were compared between groups. RESULTS: A statistically significant between group differences was recorded for body weight (BW), BCS, serum Insulin and triglyceride (TG) levels (p < 0.05). RNA-seq revealed the upregulated 154 genes and the downregulated 198 genes in obese dogs at more than 3.5-fold change compared with control animals. Hemoglobin subunits alpha- and beta-like were detected in the downregulated genes. RT-PCR analysis showed downregulation of FOLH1, ALAS2 and LOC100855540 genes, and upregulation of BCL2L15 gene, suggesting that the metabolic difference between normal and mild to moderate obesity was involved in the hemoglobin metabolism. CONCLUSIONS: This study revealed significant differences in the gene expression of BCL2L15, FOLH1, ALAS2, and hemoglobin subunits such as LOC100855540 between normal weight and mild to moderate obese dogs, which indicate that these genes may prevent the obesity in dogs and be potentially useful for diagnosis of mild to moderate obesity.

Biochemical characterization of novel lignans isolated from the wood of Taxus yunnanensis as effective stimulators for glycogen synthase kinase-3ß and the phosphorylation of basic brain proteins by the kinase in vitro.

The stimulatory and inhibitory effects of several compounds and lignans isolated from the water extract of Taxus yunnanensis on the phosphorylation of three functional brain proteins (bovine myelin basic protein (bMBP), recombinant human tau protein (rhTP) and rat collapsin response mediator protein-2 (rCRMP-2)) by glycogen synthase kinase-3ß (GSK-3ß) were quantitatively compared in vitro, using (-)-epigallocatechin-3-gallate [(-)EGCG] as a positive control. We found that (i) three selected Taxus lignans [(3S,4R)-4'-hydroxy-6,3'-dimethoxyisoflavan-4-ol,(7R)-7-hydroxytaxiresinol and tanegool] highly stimulated the autophosphorylation of GSK-3ß and the GSK-3ß-mediated phosphorylation of two basic brain proteins [bMBP (pI=11.3) and rhTP (pI=8.2)], but inhibited dose-dependently the phosphorylation of an acidic protein (rCRMP-2, pI=6.0) by the kinase; (ii) these three Taxus lignans showed binding-affinities with bMBP as well as rhTP, but had low affinities with rCRMP-2; (iii) the binding of tanegool and (7R)-7-hydroxytaxiresinol to these two basic proteins induced their novel potent phosphorylation sites for GSK-3ß; and (iv) these three Taxus lignans, but not EGCG, induced Tyr-phosphorylation of GSK-3ß in vitro. These results provided here suggest that (i) these three Taxus lignans act as novel effective activators for GSK-3ß and the GSK-3ß-mediated phosphorylation of their binding basic proteins (rhTP and bMBP); and (ii) tanegool (IC(50)=1 µM) is an effective inhibitor for the phosphorylation of rCRMP-2 by the kinase in vitro.

Biochemical characterization of epigallocatechin-3-gallate as an effective stimulator for the phosphorylation of its binding proteins by glycogen synthase kinase-3ß in vitro.

The stimulatory and inhibitory effects of epigallocatechin-3-gallate (EGCG) and its related two compounds (luteolin and quercetin) on the phosphorylation of four proteins [bovine myelin basic protein (bMBP), human recombinant tau protein (hrTP), human recombinant vimentin (hrVM) and rat collapsin response mediator protein-2 (rCRMP-2)] by glycogen synthase kinase-3ß (GSK-3ß) were comparatively determined in vitro. We found that (i) EGCG, not quercetin and luteolin, highly stimulated the GSK-3ß-mediated phosphorylation of hrTP and significantly stimulated the phosphorylation of bMBP and hrVM by the kinase; (ii) these three polyphenols inhibited dose-dependently the phosphorylation of rCRMP-2 by GSK-3ß; (iii) only EGCG significantly enhanced autophosphorylation of GSK-3ß; and (iv) EGCG had a binding-affinity with two basic proteins (bMBP and hrTP) and a low affinity with rCRMP-2 rather than hrVM in vitro. In addition, the binding of EGCG to these two basic proteins induced to highly stimulate their phosphorylation, including novel potent sites for GSK-3ß, and to significantly reduce the K(m) value and increase the V(max) value of these two substrate proteins for the kinase in vitro. These results provided here suggest that EGCG acts as an effective stimulator for the GSK-3ß-mediated phosphorylation of its binding proteins containing EGCG-inducible phosphorylation sites for the kinase in vitro.

Characterization of meFucoidan as a selective inhibitor for secretory phospholipase A2-IIA and the phosphorylation of meFucoidan-binding proteins by A-kinase in vitro.

The direct interaction of Mekabu fucoidan (meFucoidan) with four functional basic proteins (sPLA2-IIA, bFGF, histone H2B and HBV core protein) and three synthetic FGF-BP peptides (sp5, GE13 and RS6) was characterized in vitro. It was found that (i) meFucoidan inhibited dose-dependently the activity of sPLA2-IIA, but not pPLA2, through its direct binding to the enzyme; (ii) sPLA2-IIA activity was sensitive to meFucoidan rather than heparin, but significantly stimulated by sulfatide; (iii) the A-kinase-mediated phosphorylation of these basic proteins, except sPLA2-IIA, and synthetic peptides, containing potent phosphorylation sites for A-kinase, was inhibited dose-dependently by meFucoidan; and (iv) two consensus meFucoidan-binding motifs (B-B-B-B-X and B-X-B-B-X; B, basic amino acid) in these basic proteins and synthetic peptides could be overlapping to the potent phosphorylation site (B-B-X-S/T) for the kinase in vitro. These results presented here suggest that meFucoidan functions as a selective inhibitor for sPLA2-IIA and the A-kinase-mediated phosphorylation of cellular meFucoidan-binding functional basic proteins in vitro.