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To identify candidate genes that could be used as diagnostic and therapeutic targets for hepatocellular carcinoma (HCC), we searched for the genes that are overexpressed in HCC by combining representational difference analysis and microarray. Genes such as glypican-3 (GPC3), insulin-like growth factor 2, long-chain fatty-acid-coenzyme A ligase 4, farnesyl diphosphate synthase were frequently identified in our screening. Northern blot analysis with these four genes confirmed their overexpression in HCC. Among them we found that GPC3 transcript is upregulated in six out of seven cases of HCC. Immunoblot and immunohistochemical staining using polyclonal anti-GPC3 antibodies further confirmed that GPC3 protein is indeed increased in HCC tumor samples. We also found that GPC3 is secreted into culture media from cell lines derived from HCC. We conclude that GPC3 is a good molecular marker for HCC. (Cancer Sci 2003; 94: 259-262) lypican-3 (GPC3) is a member of the glypican family of heparan-sulfate proteoglycans, which are linked to the cell surface through a glycosylphosphatidylinositol anchor.1) GPC3 loss-of-function mutation in human causes type 1 Simpson-Golabi-Behmel syndrome (SGBS1), an X-linked condition characterized by pre-and postnatal overgrowth.2) GPC3 knockout mice indeed exhibited several phenotypic features of SGBS1. [3][4][5] These findings together with cell line-specific promotion of apoptosis by OCI-5/GPC3 6) suggest that GPC3 plays a negative role in cell proliferation and an apoptosis-inducing role in specific tissues.Consistent with the above idea, GPC3 expression is frequently silenced by promoter methylation in ovarian cancer cell lines, 7) rat mesothelioma cell lines and human primary tumors, 8) and breast cancer cell lines. 9) In addition, ectopic expression of GPC3 inhibited growth in some of the above cell lines, suggesting a tumor-suppressive role of GPC3. In contrast, GPC3 is known to be overexpressed in hepatocellular carcinoma, 10,11) neuroblastoma and Wilms' tumor cells.12) The role of GPC3 in these tumors is not known. It is also not known whether GPC3 protein is indeed increased in these tumors.Hepatocellular carcinoma (HCC) is one of the most common tumors worldwide and is one of the leading causes of death among cancer patients in Korea. Identification of genes that are overexpressed in HCC not only helps our understanding of tumorigenesis, but also helps to develop diagnostic and therapeutic targets. In this study, we combined representational difference analysis (RDA) 13) and microarray 14) to identify genes that are frequently overexpressed in HCC tumor samples. Since GPC3 was the most frequently obtained gene in our screening, we further evaluated it as a tumor marker for HCC. Materials and MethodsTumor samples and cell lines. HCC tumor tissues and corresponding normal liver tissues were obtained from patients (Table 1) undergoing surgery in Kyungpook National University Hospital (Daegu, Korea) with the approval of the human research review committee and the patients' consent. C...

Objective. To investigate the effector function exerted by type II collagen (CII)-stimulated T cells on rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS), and to determine their contribution to RA pathogenesis.Methods. We used enzyme-linked immunosorbent assays to measure the levels of interleukin-15 (IL-15), tumor necrosis factor ␣ (TNF␣), and IL-18 production by FLS that were cocultured with antigen-activated T cells. Likewise, we analyzed the levels of interferon-␥ (IFN␥) and IL-17 production by RA T cells coincubated with FLS. To investigate the cross-talk between CIIstimulated T cells and RA FLS, we examined the effect of using a transwell membrane to separate T cells and FLS in a culture chamber, as well as the effect of adding an antibody to block CD40 ligation.Results. The levels of IL-15, TNF␣, IFN␥, and IL-17 were all significantly increased in the serum of RA patients compared with normal control serum. Among the patients, the group with a stronger T cell proliferation response to CII showed higher levels of these

nesterified arachidonic acid (also called free AA) entering cells exogenously or released endogenously is rapidly converted to AA-CoA esters by the catalytic action of fatty acidCoA ligase (FACL), particularly by the AA-preferring FACL4.1, 2) Several lines of evidence indicated that the level of free AA in cells regulates apoptosis.3-7) More recently, it has been demonstrated that free AA induces apoptosis by activating the caspase-3 pathway, and that the induction of apoptosis by inhibitors of AA metabolism is a consequence of its accumulation.8) Moreover, overexpression of FACL4 prevented AA-induced apoptosis by reducing the level of intracellular free AA. 8)These findings, together with overexpression of FACL4 in colon adenocarcinoma, strongly suggest that the FACL4 pathway may be important in colon carcinogenesis.9) In addition, cyclooxygenase-2 (Cox-2), another AA-utilizing enzyme, is also known to be overexpressed in colon cancer.10, 11) Therefore, AA metabolism plays an important role in colon carcinogenesis. Currently, overexpression of FACL4 and Cox-2 in colon adenocarcinoma is believed to deplete free AA, thereby removing a proapoptotic signal and promoting carcinogenesis. 8,9) Hepatocellular carcinoma (HCC) is one of the most common tumors worldwide and is one of the leading causes of death among cancer patients in Korea. Identification of genes that are overexpressed in HCC is of importance to understand liver tumorigenesis and to develop diagnostic and therapeutic targets. As is seen in colon cancer, Cox-2 is known to be overexpressed in human HCC, [12][13][14] implying that AA metabolism might also be important in liver carcinogenesis. However, it is not known whether FACL4 is also overexpressed in HCC. It is known that human placenta, brain, testis, ovary, spleen, and adrenal cortex express high levels of FACL4, whereas the gastrointestinal system, including liver, expresses a very low level.2) In a recent paper, 15) we reported that FACL4 is frequently upregulated in HCC compared to chronic hepatitis, as revealed by highthroughput screening combining RDA and microarray. We think that assessing the status of FACL4 in HCC is of importance to check whether the FACL4 pathway is also involved in liver carcinogenesis. Therefore, in this study, we investigated FACL4 upregulation in tumor cells of HCC and in cell lines derived from hepatoma. Materials and MethodsTumor samples and cell lines.

Abbreviations: GSPE, grape seed proanthocyanidin extract; MIA, monosodium iodoacetate; MMP, matrix metalloproteinase; O2 -, superoxide anion; OA, osteoarthritis; OH, hydroxyl radicals; ONOO -, peroxynitrite; PWL, paw withdrawal latency; PWT, paw withdrawal threshold; ROS, reactive oxygen species; TRP, transient receptor potential; VIP, vasoactive intestinal peptide Abstract Osteoarthritis (OA) is an age-related joint disease that is characterized by degeneration of articular cartilage and chronic pain. Oxidative stress is considered one of the pathophysiological factors in the progression of OA. We investigated the effects of grape seed proanthocyanidin extract (GSPE), which is an antioxidant, on monosodium iodoacetate (MIA)-induced arthritis of the knee joint of rat, which is an animal model of human OA. GSPE (100 mg/kg or 300 mg/kg) or saline was given orally three times per week for 4 weeks after the MIA injection. Pain was measured using the paw withdrawal latency (PWL), the paw withdrawal threshold (PWT) and the hind limb weight bearing ability. Joint damage was assessed using histological and microscopic analysis and microcomputerized tomography. Matrix metalloproteinase-13 (MMP13) and nitrotyrosine were detected using immunohistochemistry. Administration of GSPE to the MIA-treated rats significantly increased the PWL and PWT and this resulted in recovery of hind paw weight distribution (P ＜ 0.05). GSPE reduced the loss of chondrocytes and proteoglycan, the production of MMP13, nitrotyrosine and IL-1β and the formation of osteophytes, and it reduced the number of subchondral bone fractures in the MIA-treated rats. These results indicate that GSPE is antinociceptive and it is protective against joint damage in the MIA-treated rat model of OA. GSPE could open up novel avenues for the treatment of OA.

White fat cells secrete adipokines that induce inflammation and obesity has been reported to be characterized by high serum levels of inflammatory cytokines such as IL-6 and TNF-α. Rheumatoid arthritis (RA) is a prototype of inflammatory arthritis, but the relationship between RA and obesity is controversial. We made an obese inflammatory arthritis model: obese collagen-induced arthritis (CIA). C57BL/6 mice were fed a 60-kcal high fat diet (HFD) from the age of 4 weeks and they were immunized twice with type II collagen (CII). After immunization, the obese CIA mice showed higher arthritis index scores and histology scores and a more increased incidence of developing arthritis than did the lean CIA mice. After treatment with CII, mixed lymphocyte reaction also showed CII-specific response more intensely in the obese CIA mice than lean CIA. The anti-CII IgG and anti-CII IgG2a levels in the sera of the obese CIA mice were higher than those of the lean CIA mice. The number of Th17 cells was higher and the IL-17 mRNA expression of the splenocytes in the obese CIA mice was higher than that of the lean CIA mice. Obese CIA mice also showed high IL-17 expression on synovium in immunohistochemistry. Although obesity may not play a pathogenic role in initiating arthritis, it could play an important role in amplifying the inflammation of arthritis through the Th1/Th17 response. The obese CIA murine model will be an important tool when we investigate the effect of several therapeutic target molecules to treat RA.

Dichloromethane (CH 2 Cl 2 ) and perchloroethylene (C 2 Cl 4 ) are chlorinated very short lived substances (Cl-VSLS) with anthropogenic sources. Recent studies highlight the increasing influence of such compounds, particularly CH 2 Cl 2 , on the stratospheric chlorine budget and therefore on ozone depletion. Here, a multiyear global-scale synthesis inversion was performed to optimize CH 2 Cl 2 (2006)(2007)(2008)(2009)(2010)(2011)(2012)(2013)(2014)(2015)(2016)(2017) and C 2 Cl 4 (2007)(2008)(2009)(2010)(2011)(2012)(2013)(2014)(2015)(2016)(2017) emissions. The approach combines long-term surface observations from global monitoring networks, output from a three-dimensional chemical transport model (TOMCAT), and novel bottom-up information on prior industry emissions. Our posterior results show an increase in global CH 2 Cl 2 emissions from 637 ± 36 Gg yr −1 in 2006 to 1,171 ± 45 Gg yr −1 in 2017, with Asian emissions accounting for 68% and 89% of these totals, respectively. In absolute terms, Asian CH 2 Cl 2 emissions increased annually by 51 Gg yr −1 over the study period, while European and North American emissions declined, indicating a continental-scale shift in emission distribution since the mid-2000s. For C 2 Cl 4 , we estimate a decrease in global emissions from 141 ± 14 Gg yr −1 in 2007 to 106 ± 12 Gg yr −1 in 2017. The time-varying posterior emissions offer significant improvements over the prior. Utilizing the posterior emissions leads to modeled tropospheric CH 2 Cl 2 and C 2 Cl 4 abundances and trends in good agreement to those observed (including independent observations to the inversion). A shorter C 2 Cl 4 lifetime, from including an uncertain Cl sink, leads to larger global C 2 Cl 4 emissions by a factor of~1.5, which in some places improves model-measurement agreement. The sensitivity of our findings to assumptions in the inversion procedure, including CH 2 Cl 2 oceanic emissions, is discussed. Plain Language SummaryThe 1987 Montreal Protocol banned production for dispersive uses of major ozone-depleting gases, such as chlorofluorocarbons, due to their role in depletion of the stratospheric ozone layer. In consequence, the ozone layer is expected to recover in coming decades, as stratospheric chlorine from banned substances slowly declines. However, chlorinated very short lived substances (Cl-VSLS), not controlled by the Montreal Protocol, represent a small, but growing, source of atmospheric chlorine that could potentially slow ozone recovery. It is thus important that the magnitude of emissions of

The potent anthropogenic perfluorocarbon greenhouse gases tetrafluoromethane (CF 4 ) and hexafluoroethane (C 2 F 6 ) are emitted to the atmosphere mainly by the aluminum and semiconductor industries. Global emissions of these perfluorocarbons (PFCs) calculated from atmospheric measurements are significantly greater than expected from reported national and industry-based emission inventories. In this study, in situ measurements of the two PFCs in the Advanced Global Atmospheric Gases Experiment network are used to show that their emission ratio varies according to the relative regional presence of these two industries, providing an industry-specific emission "signature" to apportion the observed emissions. Our results suggest that underestimated emissions from the global semiconductor industry during 1990-2010, as well as from China's aluminum industry after 2002, account for the observed differences between emissions based on atmospheric measurements and on inventories. These differences are significant despite the large uncertainties in emissions based on the methodologies used by these industries.