Hyo Jeong Kim

Hypertension and abnormal fat distribution but not insulin resistance in mice with P465L PPARγ

Peroxisome proliferator-activated receptor gamma (PPARgamma), the molecular target of a class of insulin sensitizers, regulates adipocyte differentiation and lipid metabolism. A dominant negative P467L mutation in the ligand-binding domain of PPARgamma in humans is associated with severe insulin resistance and hypertension. Homozygous mice with the equivalent P465L mutation die in utero. Heterozygous mice grow normally and have normal total adipose tissue weight. However, they have reduced interscapular brown adipose tissue and intra-abdominal fat mass, and increased extra-abdominal subcutaneous fat, compared with wild-type mice. They have normal plasma glucose levels and insulin sensitivity, and increased glucose tolerance. However, during high-fat feeding, their plasma insulin levels are mildly elevated in association with a significant increase in pancreatic islet mass. They are hypertensive, and expression of the angiotensinogen gene is increased in their subcutaneous adipose tissues. The effects of P465L on blood pressure, fat distribution, and insulin sensitivity are the same in both male and female mice regardless of diet and age. Thus the P465L mutation alone is sufficient to cause abnormal fat distribution and hypertension but not insulin resistance in mice. These results provide genetic evidence for a critical role for PPARgamma in blood pressure regulation that is not dependent on altered insulin sensitivity.

Salvianolic Acid B Exerts Vasoprotective Effects through the Modulation of Heme Oxygenase-1 and Arginase Activities

Salvia miltiorrhiza (Danshen), a traditional Chinese herbal medicine, is commonly used for the prevention and treatment of cardiovascular disorders including atherosclerosis. However, the mechanisms responsible for the vasoprotective effects of Danshen remain largely unknown. Salvianolic acid B (Sal B) represents one of the most bioactive compounds that can be extracted from the water-soluble fraction of Danshen. We investigated the effects of Danshen and Sal B on the inflammatory response in murine macrophages. Danshen and Sal B both induced the expression of heme oxygenase-1 (HO-1) and inhibited nitric oxide (NO) production and inducible NO synthase (iNOS) expression in lipopolysaccharide (LPS)-activated RAW 264.7 cells. Inhibition of HO activity using Sn-protoporphyrin-IX (SnPP) abolished the inhibitory effect of Sal B on NO production and iNOS expression. Sal B increased macrophage arginase activity in a dose-dependent manner and diminished LPS-inducible tumor necrosis factor-α production. These effects were also reversed by SnPP. These data suggest that HO-1 expression plays an intermediary role in the anti-inflammatory effects of Sal B. In contrast to the observations in macrophages, Sal B dose-dependently inhibited arginase activity in murine liver, kidney, and vascular tissue. Furthermore, Sal B increased NO production in isolated mouse aortas through the inhibition of arginase activity and reduction of reactive oxygen species production. We conclude that Sal B improves vascular function by inhibiting inflammatory responses and promoting endothelium-dependent vasodilation. Taken together, we suggest that Sal B may represent a potent candidate therapeutic for the treatment of cardiovascular diseases associated with endothelial dysfunction.

Stability of the LATS2 Tumor Suppressor Gene Is Regulated by Tristetraprolin

LATS2 is a tumor suppressor gene implicated in the control of cell growth and the cell cycle. Here, we investigated the post-transcriptional regulation of LATS2 expression by tristetraprolin (TTP). Our results show that the expression level of LATS2 is inversely correlated with TTP expression in human cancer cell lines. Overexpression of TTP reduced the expression level of LATS2. Conversely, treatment with small interfering RNA against TTP increased the expression level of LATS2 through stabilization of LATS2 mRNA and suppressed the proliferation of A549 human lung cancer cells. LATS2 mRNA contains AU-rich elements (AREs) within the 3′-untranslated region, and TTP destabilized a luciferase mRNA containing LATS2 ARE. In addition, RNA electrophoretic mobility shift assay revealed that TTP directly bound to the ARE of LATS2 mRNA. These results establish LATS2 mRNA as a physiological target of TTP and suggest the possibility that TTP controls cell growth through regulation of LATS2 mRNA stability.

Tristetraprolin Mediates Anti-inflammatory Effects of Nicotine in Lipopolysaccharide-stimulated Macrophages

Nicotine inhibits the release of TNF-α from macrophage through activation of STAT3. Tristetraprolin (TTP) is known to destabilize pro-inflammatory transcripts containing AU-rich elements (ARE) in 3′-untranslated region (3′-UTR). Here we show that in LPS-stimulated human macrophages the anti-inflammatory action of nicotine is mediated by TTP. Nicotine induced activation of STAT3 enhanced STAT3 binding to the TTP promoter, increased TTP promoter activity, and increased TTP expression resulting in the suppression of LPS-stimulated TNF-α production. Overexpression of a dominant negative mutant of STAT3 (R382W) or down-regulation of STAT3 by siRNA abolished nicotine-induced TTP expression and suppression of LPS-stimulated TNF-α production. Nicotine enhanced the decay of TNF-α mRNA and decreased luciferase expression of a TNF-α 3′-UTR reporter plasmid in U937 cells. However, siRNA to TTP abrogated these effects of nicotine. In this experiment, we are reporting for the first time the involvement of TTP in the cholinergic anti-inflammatory cascade consisting of nicotine-STAT3-TTP-dampening inflammation.

Resveratrol Induces Hepatic Mitochondrial Biogenesis Through the Sequential Activation of Nitric Oxide and Carbon Monoxide Production

AIMSnNitric oxide (NO) can induce mitochondrial biogenesis in cultured cells, through increased guanosine 3,5-monophosphate (cGMP), and activation of peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α). We sought to determine the role of NO, heme oxygenase-1 (HO-1), and its reaction product (carbon monoxide [CO]) in the induction of mitochondrial biogenesis by the natural antioxidant resveratrol.nnnRESULTSnS-nitroso-N-acetylpenicillamine (SNAP), an NO donor, induced mitochondrial biogenesis in HepG2 hepatoma cells, and in vivo, through stimulation of PGC-1α. NO-induced mitochondrial biogenesis required cGMP, and was mimicked by the cGMP analogue (8-bromoguanosine 3,5-cyclic monophosphate [8-Br-cGMP]). Activation of mitochondrial biogenesis by SNAP required HO-1, as it could be reversed by genetic interference of HO-1; and by treatment with the HO inhibitor tin-protoporphyrin-IX (SnPP) in vitro and in vivo. Cobalt protoporphyrin (CoPP)-IX, an HO-1 inducing agent, stimulated mitochondrial biogenesis in HepG2 cells, which could be reversed by the CO scavenger hemoglobin. Application of CO, using the CO-releasing molecule-3 (CORM-3), stimulated mitochondrial biogenesis in HepG2 cells, in a cGMP-dependent manner. Both CoPP and CORM-3-induced mitochondrial biogenesis required NF-E2-related factor-2 (Nrf2) activation and phosphorylation of Akt. The natural antioxidant resveratrol induced mitochondrial biogenesis in HepG2 cells, in a manner dependent on NO biosynthesis, cGMP synthesis, Nrf2-dependent HO-1 activation, and endogenous CO production. Furthermore, resveratrol preserved mitochondrial biogenesis during lipopolysaccharides-induced hepatic inflammation in vivo.nnnINNOVATION AND CONCLUSIONSnThe complex interplay between endogenous NO and CO production may underlie the mechanism by which natural antioxidants induce mitochondrial biogenesis. Strategies aimed at improving mitochondrial biogenesis may be used as therapeutics for the treatment of diseases involving mitochondrial dysfunction.

Casein kinase 2 regulates the mRNA-destabilizing activity of tristetraprolin.

Tristetraprolin (TTP) is an AU-rich element-binding protein that regulates mRNA stability. We previously showed that TTP acts as a negative regulator of VEGF gene expression in colon cancer cells. The p38 MAPK pathway is known to suppress the TTP activity. However, until now the signaling pathway to enhance TTP function is not well known. Here, we show that casein kinase 2 (CK2) enhances the TTP function in the regulation of the VEGF expression in colon cancer cells. CK2 increased TTP protein levels and enhanced VEGF mRNA decaying activity of TTP. TTP was not a direct target of CK2. Instead, CK2 increased the phosphorylation of MKP-1, which led to a decrease in the phosphorylation of p38 MAPK. Inhibition of MKP-1 by siRNA attenuated the increase in TTP function and the decrease of p38 phosphorylation induced by CK2α overexpression. TGF-β1 increased the expressions of CK2 and TTP and the TTP function. The siRNA against CK2α or TTP reversed TGF-β1-induced increases in the expression of CK2 and TTP and the TTP function. Our data suggest that CK2 enhances the protein level and activity of TTP via the modulation of the MKP-1-p38 MAPK signaling pathway and that TGF-β1 enhances the activity of CK2.

Microfluidic Separation of a Soluble Substance Using Transverse Diffusion in a Layered Flow

This paper presents a practical flow-through method to separate anisole and ethyl phenylacetate, respectively, from a polystyrene mixture. The microfluidic separation uses different diffusive dynamics of the substances transverse to the lamination flow formed in a microchannel. The effect of inlet flow rates and ambient temperature on separation is examined. Additionally, the possibility of the separation of the light substance from the mixture with different molecular weight is shown numerically and experimentally. The separation efficiency is explained by the facts that the relaxation time depends on the inlet flow rate and that the diffusivity depends on the ambient temperature. This method can be applied to separate monomers from aggregates.