Although the study did not meet its primary end point, the trend toward improved OS with FOLFOX4, along with increased PFS and RR, suggests that this regimen may confer some benefit to Asian patients, but an OS benefit cannot be concluded from these data.

Abstract-Toll-like receptor (TLR)4 recognizes microbial pathogens, such as lipopolysaccharide, and mediates lipopolysaccharide-induced proinflammatory cytokine secretion, as well as microbial uptake by macrophages. In addition to exogenous pathogens, TLR4 recognizes modified self, such as minimally oxidized low-density lipoprotein (mmLDL). Here we report that mmLDL and its active components, cholesteryl ester hydroperoxides, induce TLR4-dependent fluid phase uptake typical of macropinocytosis. We show that mmLDL induced recruitment of spleen tyrosine kinase (Syk) to a TLR4 signaling complex, TLR4 phosphorylation, activation of a Vav1-Ras-Raf-MEK-ERK1/2 signaling cascade, phosphorylation of paxillin, and activation of Rac, Cdc42, and Rho. These mmLDL-induced and TLR4-and Syk-dependent signaling events and cytoskeletal rearrangements lead to enhanced uptake of small molecules, dextran, and, most importantly, both native and oxidized LDL, resulting in intracellular lipid accumulation. An intravenous injection of fluorescently labeled mmLDL in wild-type mice resulted in its rapid accumulation in circulating monocytes, which was significantly attenuated in TLR4-deficient mice. These data describe a novel mechanism leading to enhanced lipoprotein uptake in macrophages that would contribute to foam cell formation and atherosclerosis. These data also suggest that cholesteryl ester hydroperoxides are an endogenous ligand for TLR4. Because TLR4 is highly expressed on the surface of circulating monocytes in patients with chronic inflammatory conditions, and cholesteryl ester hydroperoxides are present in plasma, lipid uptake by monocytes in circulation may contribute to the pathological roles of monocytes in chronic inflammatory diseases. T oll-like receptors (TLRs) are pattern recognition receptors (PRRs) that sense the presence of numerous pathogen-associated molecular patterns (PAMPs). 1 Activation of PRRs has been widely implicated in signaling mechanisms that contribute to chronic inflammatory diseases, including atherosclerosis. Although PRRs were originally postulated to recognize only exogenous pathogens, they are now increasingly documented to respond to endogenous modified self, such as modified and/or oxidized low-density lipoprotein (LDL). We have postulated that such modified LDL could become endogenous PAMPs and initiate lowgrade, but sustained, PRR-mediated inflammation and other immune responses. 2 TLR4 and TLR2 deficiency, as well as a deficiency in MyD88, a signaling adaptor molecule for TLR4 and other TLRs, result in reduced atherosclerosis in apoE Ϫ/Ϫ or LDLR Ϫ/Ϫ mice fed a high-fat diet. [3][4][5] Some human studies, though not all, have shown that inactivating polymorphisms in TLR4 are associated with decreased cardiovascular disease risk. 6 Although these data support a general understanding that TLRs regulate inflammation in atherosclerosis, specific mechanisms are poorly understood.Excessive lipoprotein accumulation by macrophages and the formation of lipid-loaded foam cells is a rate-limiting pro...

Abstract-Oxidative modification of low-density lipoprotein (LDL) plays a causative role in the development of atherosclerosis. In this study, we demonstrate that minimally oxidized LDL (mmLDL) stimulates intracellular reactive oxygen species (ROS) generation in macrophages through NADPH oxidase 2 (gp91phox/Nox2), which, in turn, induces production of RANTES and migration of smooth muscle cells. Peritoneal macrophages from gp91phox/Nox2 Ϫ/Ϫ mice or J774 macrophages in which Nox2 was knocked down by small interfering RNA failed to generate ROS in response to mmLDL. Because mmLDL-induced cytoskeletal changes were dependent on Toll-like receptor (TLR)4, we analyzed ROS generation in peritoneal macrophages from wild-type, TLR4 Ϫ/Ϫ , or MyD88 Ϫ/Ϫ mice and found that mmLDLmediated ROS was generated in a TLR4-dependent, but MyD88-independent, manner. Furthermore, we found that ROS generation required the recruitment and activation of spleen tyrosine kinase (Syk) and that mmLDL also induced phospholipase PLC␥1 phosphorylation and protein kinase C membrane translocation. Importantly, the phospholipase C␥1 phosphorylation was reduced in J774 cells expressing Syk-specific short hairpin RNA. Nox2 modulated mmLDL activation of macrophages by regulating the expression of proinflammatory cytokines interleukin-1␤, interleukin-6, and RANTES. We showed that purified RANTES was able to stimulate migration of mouse aortic smooth muscle cells and addition of neutralizing antibody against RANTES abolished the migration of mouse aortic smooth muscle cells stimulated by mmLDL-stimulated macrophages. These results suggest that mmLDL induces generation of ROS through sequential activation of TLR4, Syk, phospholipase C␥1, protein kinase C, and gp91phox/Nox2 and thereby stimulates expression of proinflammatory cytokines. These data help explain mechanisms by which endogenous ligands, such as mmLDL, can induce TLR4-dependent, proatherogenic activation of macrophages. (Circ Res. 2009;104:210-218.) Key Words: minimally oxidized LDL Ⅲ reactive oxygen species Ⅲ NADPH oxidase 2 Ⅲ RANTES Ⅲ atherosclerosis R eactive oxygen species (ROS), including hydrogen peroxide and superoxide anion, are generally considered cytotoxic. 1 However, in recent years, many reports have demonstrated that intracellular ROS, produced in mammalian cells in response to the activation of various receptors, also serve as important second messengers in cell signaling. 1 The major source of ROS in phagocytes is an NADPH oxidase complex. 2 It is composed of 6 protein components: 2 transmembrane flavocytochrome b proteins (gp91phox and p22phox) and 4 cytosolic proteins (p47phox, p67phox, p40phox, and Rac). The catalytic protein in this complex is gp91phox, recently renamed as Nox2. The NH 3 -terminal region of gp91phox/Nox2 consists of transmembrane domains that bind 2 heme groups, and the COOH-terminal region contains NADPH/FAD binding sites. The cytosolic component of the complex, p47phox, plays a critical role in an assembly of the whole NADPH oxidase complex on activation o...

A lexicon for describing green tea was developed using descriptive analysis methods. A highly trained, descriptive sensory panel identified, defined and referenced 31 flavor attributes for green tea. One-hundred and thirty-eight green tea samples from nine countries and Vietnam -were selected to represent a wide range of green teas. Attributes could be categorized as "Green" (asparagus, beany, Brussels sprout, celery, parsley, spinach, green beans, green herb-like); "Brown" (ashy/sooty, brown spice, burnt/scorched, nutty, tobacco); "Fruity/ Floral" (fruity, floral/perfumy, citrus, fermented); "Mouthfeel" (astringent, tooth-etching); "Basic Tastes" (overall sweet, bitter); and other attributes (almond, animalic, grain, musty/new leather, mint, seaweed, straw-like). Some attributes, such as green, brown, bitter, astringent and tooth-etching, were found in most samples, but many attributes were found in only a few samples. Green tea processors, food industry, researchers and consumers will benefit from this lexicon with precise definitions and references that reliably differentiate and characterize the sensory attributes of green teas. PRACTICAL APPLICATIONSGreen tea (and white tea) processors, food industrialists, researchers and consumers will benefit from this lexicon with precise definitions and references that reliably differentiate and characterize the sensory attributes of green tea.

Quorum sensing (QS) controls certain behaviors of bacteria in response to population density. In Gram-negative bacteria, QS is often mediated by N-acyl-L-homoserine lactones (acyl-HSLs). Because QS influences the virulence of many pathogenic bacteria, synthetic inhibitors of acyl-HSL synthases might be useful therapeutically for controlling pathogens. However, rational design of a potent QS antagonist has been thwarted by the lack of information concerning the binding interactions between acyl-HSL synthases and their ligands. In the Gram-negative bacterium Burkholderia glumae, QS controls virulence, motility, and protein secretion and is mediated by the binding of N-octanoyl-L-HSL (C8-HSL) to its cognate receptor, TofR. C8-HSL is synthesized by the acyl-HSL synthase TofI. In this study, we characterized two previously unknown QS inhibitors identified in a focused library of acyl-HSL analogs. Our functional and X-ray crystal structure analyses show that the first inhibitor, J8-C8, binds to TofI, occupying the binding site for the acyl chain of the TofI cognate substrate, acylated acyl-carrier protein.Moreover, the reaction byproduct, 5′-methylthioadenosine, independently binds to the binding site for a second substrate, Sadenosyl-L-methionine. Closer inspection of the mode of J8-C8 binding to TofI provides a likely molecular basis for the various substrate specificities of acyl-HSL synthases. The second inhibitor, E9C-3oxoC6, competitively inhibits C8-HSL binding to TofR. Our analysis of the binding of an inhibitor and a reaction byproduct to an acyl-HSL synthase may facilitate the design of a new class of QS-inhibiting therapeutic agents. Q uorum sensing (QS) is an intercellular signaling process that mediates certain behaviors of bacteria (including bioluminescence, biofilm formation, motility, and virulence factor production) in response to the bacterial cell population density (1-3). In Gram-negative bacteria, QS is often mediated by Nacyl-L-homoserine lactones (acyl-HSLs), which are synthesized by the LuxI family of acyl-HSL synthases from S-adenosyl-Lmethionine (SAM) and acylated acyl-carrier protein (acyl-ACP), with the release of holo-ACP and 5′-methylthioadenosine (MTA) as byproducts (SI Appendix, Fig. S1A) (4, 5). Compounds of the acyl-HSL class share a homoserine lactone ring moiety, but the acyl chains conjugated to the ring via an amide bond vary in length, oxidation state at C3, and amount of saturation (SI Appendix, Fig. S1A). The recent finding that p-coumarate is an alternative substrate for acyl-ACP has extended the known range of possible acyl-HSL substrates (6). On the other hand, the acyl-HSL receptor is a transcriptional regulator that controls the expression of target genes in response to acyl-HSL binding (1-3).Among the hundreds of genes regulated by QS, the most widely studied genes are those related to virulence; these genes are of particular interest because QS disruption is being investigated as a strategy for controlling virulent pathogens (7-9). QS inhibitors can act by suppressing a...

PURPOSE Adjuvant chemotherapy after D2 gastrectomy is standard for resectable locally advanced gastric cancer (LAGC) in Asia. Based on positive findings for perioperative chemotherapy in European phase III studies, the phase III PRODIGY study (ClinicalTrials.gov identifier: NCT01515748 ) investigated whether neoadjuvant docetaxel, oxaliplatin, and S-1 (DOS) followed by surgery and adjuvant S-1 could improve outcomes versus standard treatment in Korean patients with resectable LAGC. PATIENTS AND METHODS Patients 20-75 years of age, with Eastern Cooperative Oncology Group performance status 0-1, and with histologically confirmed primary gastric or gastroesophageal junction adenocarcinoma (clinical TNM staging: T2-3N+ or T4Nany) were randomly assigned to D2 surgery followed by adjuvant S-1 (40-60 mg orally twice a day, days 1-28 q6w for eight cycles; SC group) or neoadjuvant DOS (docetaxel 50 mg/m2, oxaliplatin 100 mg/m2 intravenously day 1, S-1 40 mg/m2 orally twice a day, days 1-14 q3w for three cycles) before D2 surgery, followed by adjuvant S-1 (CSC group). The primary objective was progression-free survival (PFS) with CSC versus SC. Two sensitivity analyses were performed: intent-to-treat and landmark PFS analysis. RESULTS Between January 18, 2012, and January 2, 2017, 266 patients were randomly assigned to CSC and 264 to SC at 18 Korean study sites; 238 and 246 patients, respectively, were treated (full analysis set). Follow-up was ongoing in 176 patients at data cutoff (January 21, 2019; median follow-up 38.6 months [interquartile range, 23.5-62.1]). CSC improved PFS versus SC (adjusted hazard ratio, 0.70; 95% CI, 0.52 to 0.95; stratified log-rank P = .023). Sensitivity analyses confirmed these findings. Treatments were well tolerated. Two grade 5 adverse events (febrile neutropenia and dyspnea) occurred during neoadjuvant treatment. CONCLUSION PRODIGY showed that neoadjuvant DOS chemotherapy, as part of perioperative chemotherapy, is effective and tolerable in Korean patients with LAGC.

Glucose uptake and glycolytic metabolism are enhanced in cancer cells compared to normal cells and tissues. Increased expression of glucose transporter 1 (GLUT1) has been reported in human malignant cells. The aim of this study is to determine the expression of the facilitative glucose transporter protein GLUT1 in human breast carcinomas and a possible correlation between GLUT1 expression and clinical outcome including disease-free or overall survival. One hundred consecutive formalin-fixed, paraffin-embedded sections of invasive breast carcinomas were evaluated by means of immunohistochemical staining of GLUT1. Forty-seven (47%) of 100 breast carcinomas showed positive staining for GLUT1. Expression of GLUT1 correlated significantly with nuclear grade (P < < < <0.001), estrogen receptor status (P = = = =0.002), and progesterone receptor statusThe mean disease-free survival periods of GLUT1-positive and -negative patients were 47 ± ± ± ±2.4 months and 54.3± ± ± ±1.3 months, respectively (P = = = =0.017). The mean overall survival periods of GLUT1-positive and -negative patients were 48.7 ± ± ± ±2.2 and 56.1 ± ± ± ±1.3 months, respectively (P = = = =0.043).In the multivariate analysis, disease-free survival correlated significantly with GLUT1, tumor size, and lymph node involvement (P = = = =0.043, P = = = =0.014, and P = = = =0.045, respectively). In analysis of overall survival, however, lymph node involvement, tumor size, and nuclear grade were statistically significant (P = = = =0.024, P = = = =0.023, and P = = = =0.003, respectively). Our data suggest that absence of GLUT1 expression significantly increases disease-free survival. These findings demonstrate that GLUT1 expression in breast carcinoma can be a marker of aggressive biological behavior and identifies a worse prognosis in breast carcinoma patients.

Hutchinson-Gilford progeria syndrome (HGPS) is a rare autosomal dominant genetic disease that is caused by a silent mutation of the LMNA gene encoding lamins A and C (lamin A/C). The G608G mutation generates a more accessible splicing donor site than does WT and produces an alternatively spliced product of LMNA called progerin, which is also expressed in normal aged cells. In this study, we determined that progerin binds directly to lamin A/C and induces profound nuclear aberrations. Given this observation, we performed a random screening of a chemical library and identified 3 compounds (JH1, JH4, and JH13) that efficiently block progerin-lamin A/C binding. These 3 chemicals, particularly JH4, alleviated nuclear deformation and reversed senescence markers characteristic of HGPS cells, including growth arrest and senescence-associated β-gal (SA-β-gal) activity. We then used microarray-based analysis to demonstrate that JH4 is able to rescue defects of cell-cycle progression in both HGPS and aged cells. Furthermore, administration of JH4 to LmnaG609G/G609G-mutant mice, which phenocopy human HGPS, resulted in a marked improvement of several progeria phenotypes and an extended lifespan. Together, these findings indicate that specific inhibitors with the ability to block pathological progerin-lamin A/C binding may represent a promising strategy for improving lifespan and health in both HGPS and normal aging.