Objective While nonsteroidal antiinflammatory drugs (NSAIDs) are commonly used in rheumatology, they can cause major toxicity. Improving the risk/benefit ratio requires a more precise understanding of risk. This study was undertaken to derive and validate a risk score for major toxicity among NSAID users enrolled in a randomized controlled trial. Methods Patients enrolled in a randomized controlled trial who had known cardiovascular disease or risk factors as well as osteoarthritis or rheumatoid arthritis were divided into derivation and validation cohorts. Patients were randomized to receive celecoxib, naproxen, or ibuprofen at typical dosages. The risk score was designed to predict the 1‐year occurrence of major toxicity among NSAID users, including major adverse cardiovascular events, acute kidney injury, significant gastrointestinal events, and mortality. Variables significantly associated with major toxicity were candidates for inclusion in the final regression model. After derived models were found to have a similar model fit in the validation set, the cohorts were combined, allowing calculation of a risk score. Results In the derivation cohort, significant variables included age, male sex, history of cardiovascular disease, hypertension, diabetes mellitus, tobacco use, statin use, elevated serum creatinine level, hematocrit level, and type of arthritis. The C‐index was 0.73 in the validation cohort and 0.71 in the total cohort; the model was well calibrated. Of the total population with complete data (n = 23,735), 1,080 participants (4.6%) had a predicted 1‐year risk of major toxicity of <1%, 16,273 (68.6%) had a predicted risk of 1–4%, and 6,382 (26.9%) had a predicted risk of >4%. Conclusion The risk score accurately categorizes the 1‐year risk of major toxicity among NSAID users and may be useful in identifying patients who can safely use these agents.

Glycine amidinotransferase (AGAT or GATM), guanidinoacetate methyltransferase (GAMT) and creatine transporter (CT1) are three proteins involved in the synthesis and uptake of creatine. The expression patterns of these three genes were examined in zebrafish embryos by whole mount in situ hybridization followed by histological sectioning. Expression of agat first appeared in the yolk syncytial layer (YSL) at the gastrula stage and was progressively up regulated during gastrulation. As development proceeds, agat was expressed in the mature somites during the segmentation stage and in the liver at 48 hpf. gamt showed a similar expression pattern to that of agat during embryogenesis. It was first detected in the center of the yolk from the cleavage to the gastrula stage. At the bud stage, its expression shifted to the YSL. gamt was also transiently expressed in the mature somites from 16 hpf to 24 hpf and became strongly expressed in the liver and in epithelial cells of the gut at 48 hpf. ct1 was initially uniformly expressed from the cleavage to the early segmentation stage; it was then strongly expressed in all the somites till 30 hpf and in the gut of 48 hpf embryos. However, ct1 transcripts also appeared in the central nervous system during the segmentation stage, but not in the YSL, the yolk or the liver. Our data reveal for the first time distinct and unique patterns of expression of the creatine metabolism genes agat, gamt and ct1 during zebrafish embryogenesis. KEY WORDS: agat, gamt, ct1, zebrafish, developmental expressionThe creatine metabolism plays a crucial role for keeping the normal life of vertebrates. Endogenous creatine is synthesized by a two-step mechanism involving two enzymes: glycine amidinotransferase (AGAT or GATM) and guanidinoacetate methyltransferase (GAMT). Creatine is taken up by cells through CT1, a specific creatine transporter (Wyss and KaddurahDaouk, 2000). Defects of AGAT, GAMT and CT1 result in three kinds of creatine deficiency syndromes (CDS) occurring mostly in children (Schulze, 2003). The common clinical feature of three CDS is developmental delay/regression, mental retardation and severe disturbance of their expressive and cognitive speech (van der . The biochemical characteristics of CDS include severe depletion of creatine/phosphocreatine in the brain, as well as changes in creatine and creatinine concentrations in body fluids , Cecil et al., 2001, Schulze, 2003, Stromberger et al., 2003, Almeida et al., 2004, Sykut-Cegielska et al., 2004. GAMT deficiency is characterized by accumulation of guanidinoacetic acid in brain and body fluids and shows intrac- Abbreviations used in this paper: AGAT (or GATM), glycine amidinotransferase; GAMT, guanidinoacetate methyltransferase; CDS, creatine deficiency syndromes; CT1, creatine transporter 1; YSL, yolk syncytial layer. al., 2004). GAMT and AGAT deficiency have autosomal-recessive traits, whereas the CT1 defect is an X-linked disorder (Mancini et al., 2005. Treatment with oral creatine supplementation is in part successful ...

Aim: The present study examined the differential expression of proteins in HuH‐7 cells and HuH‐7 cells harboring in vitro‐transcribed full‐length hepatitis C virus 1b RNA (HuH‐7‐HCV), and elucidated the cellular responses to HCV replication. Methods: The protein profiles of matched pairs of HuH‐7‐HCV cells and HuH‐7 mock cells were analyzed by 2‐D electrophoresis (2DE). Solubilized proteins were separated in the first dimension by isoelectric focusing, and by 12.5% SDS‐PAGE in the second dimension. The differential protein expression was analyzed by use of image analysis software to identify candidates for HCV infection‐associated proteins. Results: In total, 29 protein spots showed increases and 25 protein spots showed decreases in signal in HuH‐7‐HCV cell 2DE profiles as compared with HuH‐7 mock cells. In the next step, the 10 spots showing the greatest increase and the 10 spots showing the greatest decrease were excised from gels and the proteins present were identified by Matrix‐Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometer (MALDI‐TOF MS) or MALDI‐TOF/TOF MS. In total, 13 proteins were identified successfully. The potential significance of the differential expression due to HCV replication was discussed. Conclusion: Our study identifies changes in the proteome of HuH‐7 cells in the presence of HCV replication and yields information of the mechanism of HCV pathogenesis. These results will be useful for the identification of HCV infection‐associated proteins that could be molecular targets for treatment.

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