Prostaglandins biosynthesis and nitric oxide production have been implicated in the process of carcinogenesis and inflammation. In this study, we investigated the effect of various flavonoids and (-)-epigallocatechin-3-gallate on the activities of inducible cyclooxygenase (COX-2) and inducible nitric oxide synthase (iNOS) in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages. Apigenin, genistein and kaempferol were markedly active inhibitors of transcriptional activation of COX-2, with IC(50) < 15 microM. In addition, apigenin and kaempferol were also markedly active inhibitors of transcriptional activation of iNOS, with IC(50) < 15 microM. Of those compounds tested, apigenin was the most potent inhibitor of transcriptional activation of both COX-2 and iNOS. Western and northern blot analyses demonstrated that apigenin significantly blocked protein and mRNA expression of COX-2 and iNOS in LPS-activated macrophages. Transient transfection experiments showed that LPS caused an approximately 4-fold increase in both COX-2 and iNOS promoter activities, these increments were suppressed by apigenin. Moreover, electrophoretic mobility shift assay (EMSA) experiments indicated that apigenin blocked the LPS-induced activation of nuclear factor-kB (NF-kB). The inhibition of NF-kB activation occurs through the prevention of inhibitor kB (IkB) degradation. Transient transfection experiments also showed that apigenin inhibited NF-kB-dependent transcriptional activity. Finally, we showed that apigenin could inhibit the IkB kinase activity induced by LPS or interferon-gamma. The results of further studies suggest that suppression of transcriptional activation of COX-2 and iNOS by apigenin might mainly be mediated through inhibition of IkB kinase activity. This study suggests that modulation of COX-2 and iNOS by apigenin and related flavonoids may be important in the prevention of carcinogenesis and inflammation.

The oxysterol receptors LXR (liver X receptor)-␣ and LXR␤ are nuclear receptors that play a key role in regulation of cholesterol and fatty acid metabolism. We found that LXRs also play a significant role in glucose metabolism. Treatment of diabetic rodents with the LXR agonist, T0901317, resulted in dramatic reduction of plasma glucose. In insulin-resistant Zucker (fa/fa) rats, T0901317 significantly improved insulin sensitivity. Activation of LXR did not induce robust adipogenesis but rather inhibited the expression of several genes involved in hepatic gluconeogenesis, including phosphoenolpyruvate carboxykinase (PEPCK). Hepatic glucose output was dramatically reduced as a result of this regulation. Nuclear run-on studies indicated that transcriptional repression was primarily responsible for the inhibition of PEPCK by the LXR agonist. In addition, we show that the regulation of the liver gluconeogenic pathway by LXR agonists was a direct effect on hepatocytes. These data not only suggest that LXRs are novel targets for diabetes but also reveal an unanticipated role for these receptors, further linking lipid and glucose metabolism.Type II diabetes mellitus is a prevalent metabolic disease in developed countries, with insufficient therapies for treatment and prevention (1, 2). Studies in recent years have suggested that nuclear receptors are intimately linked to the pathophysiology of diabetes. The antidiabetic thiazolidinediones have been identified as ligands of proxisome proliferator-activated receptor ␥ (PPAR␥) 1 (3, 4). Retinoid X receptor (RXR) ligands have also been shown to lower plasma glucose levels in rodent diabetic models (3-5).Originally identified as orphan members of the nuclear receptor superfamily, liver X receptors exist as two isoforms, LXR␣ and LXR␤. The two isoforms display distinct patterns of expression with LXR␣ being primarily expressed in liver, intestine, and kidney, whereas LXR␤ is expressed ubiquitously (6). Oxysterols were identified as the putative physiological ligands for the LXRs (7), and additional studies have demonstrated that these receptors act as sensors for these cholesterol metabolites and are essential components of a physiological feedback loop regulating cholesterol metabolism and transport (8). Consistent with their role in regulation of these metabolic pathways, several LXR-regulated genes involved in lipid metabolism and cholesterol transport have been identified including ABCA1, ABCG1, ABCG5, ABCG8, ApoE, CETP, Cyp7a, LPL, SREBP1c, and FAS (8 -14).As a result of the close relationship between lipid and carbohydrate metabolism, we examined the potential role LXRs may play in glucose homeostasis by using a specific LXR agonist, T0901317, (11) in rodent models of diabetes. Our findings indicated that T0901317 dose-dependently lowered plasma glucose level in both db/db and Zucker diabetic fatty (ZDF) rat models. In the fa/fa insulin-resistant rat model, T0901317 significantly improved insulin sensitivity. Examination of the liver gluconeogenesis pathway revealed dra...

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Carnosol is a naturally occurring phytopolyphenol found in rosemary. Carnosol functions as antioxidant and anticarcinogen. In the present study, we compared the antioxidant activity of carnosol and other compounds extracted from rosemary. Carnosol showed potent antioxidative activity in alpha,alpha-diphenyl-beta-picrylhydrazyl (DPPH) free radicals scavenge and DNA protection from Fenton reaction. High concentrations of nitric oxide (NO) are produced by inducible NO synthase (iNOS) in inflammation and multiple stages of carcinogenesis. Treatment of mouse macrophage RAW 264.7 cell line with carnosol markly reduced lipopolysaccharide (LPS)-stimulated NO production in a concentration-related manner with an IC50 of 9.4 microM; but other tested compounds had slight effects. Western blot, reverse transcription-polymerase chain reaction, and northern blot analyses demonstrated that carnosol decreased LPS-induced iNOS mRNA and protein expression. Carnosol treatment showed reduction of nuclear factor-kappaB (NF-kappaB) subunits translocation and NF-kappaB DNA binding activity in activated macrophages. Carnosol also showed inhibition of iNOS and NF-kappaB promoter activity in transient transfection assay. These activities were referred to down-regulation of inhibitor kappaB (IkappaB) kinase (IKK) activity by carnosol (5 microM), thus inhibited LPS-induced phosphorylation as well as degradation of IkappaBalpha. Carnosol also inhibited LPS-induced p38 and p44/42 mitogen-activated protein kinase (MAPK) activation at a higher concentration (20 microM). These results suggest that carnosol suppresses the NO production and iNOS gene expression by inhibiting NF-kappaB activation, and provide possible mechanisms for its anti-inflammatory and chemopreventive action.

Tea polyphenols are known to inhibit a wide variety of enzymatic activities associated with cell proliferation and tumor progression. The molecular mechanisms of antiproliferation are remained to be elucidated. In this study, we investigated the effects of the major tea polyphenol (-)-epigallocatechin gallate (EGCG) on the proliferation of human epidermoid carcinoma cell line, A431. Using a [3H]thymidine incorporation assay, EGCG could significantly inhibit the DNA synthesis of A431 cells. In vitro assay, EGCG strongly inhibited the protein tyrosine kinase (PTK) activities of EGF-R, PDGF-R, and FGF-R, and exhibited an IC50 value of 0.5-1 microgram/ml. But EGCG scarcely inhibited the protein kinase activities of pp60v-src, PKC, and PKA (IC50 > 10 micrograms/ml). In an in vivo assay, EGCG could reduce the autophosphorylation level of EGF-R by EGF. Phosphoamino acid analysis of the EGF-R revealed that EGCG inhibited the EGF-stimulated increase in phosphotyrosine level in A431 cells. In addition, we showed that EGCG blocked EGF binding to its receptor. The results of further studies suggested that the inhibition of proliferation and suppression of the EGF signaling by EGCG might mainly mediate dose-dependent blocking of ligand binding to its receptor, and subsequently through inhibition of EGF-R kinase activity.