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Background Proton pump inhibitors (PPIs) are common and widely used for gastrointestinal-related disorders. Lansoprazole is one of PPIs with potential benefits of anti-inflammation, reduced oxidative stress, and anti-diabetes. The aims of this study are to determine whether lansoprazole imparts differential risk of type 2 diabetes as compared with other PPIs. Methods A population-based retrospective cohort study was conducted using the National Health Insurance Research Database in Taiwan. Patients who received lansoprazole more than 90 days and without records of use of other PPIs between January 1, 2000 and December 31, 2005 (the exposure period) were considered as the exposed cohort (n = 1668). In comparison, patients who received other PPIs more than 90 days and without use of lansoprazole in the exposure period were treated as the comparison cohort (n = 3336).The primary outcome was the new-onset of type 2 diabetes mellitus (T2DM). The association between use of lansoprazole and the risk of T2DM was determined by hazard ratios (HRs) and 95% confidence intervals (CIs) derived from multivariable Cox proportional hazards models. Results The lansoprazole cohort showed a significantly reduced risk of T2DM with an adjusted HR of 0.65 (95% CI 0.56–0.76). Interestingly, the inverse association between use of lansoprazole and risk of T2DM was observed in both genders and in various age groups. Conclusion The present study findings suggest that lansoprazole was associated with a reduced risk of T2DM compared with other PPIs. Further studies are needed to determine the clinical implications of the present study.

Methyl gallate is a polyphenolic compound found in many plants, and its antioxidant, antitumor, antibacterial, and anti-inflammatory effects have been extensively studied. More recently, antidepressant-like effects of methyl gallate have been demonstrated in some studies. In the present study, we examined the effects of methyl gallate on melanogenesis, including the tyrosinase inhibitory effect, the melanin content, and the molecular signaling pathways involved in this inhibition. The results showed that methyl gallate inhibited tyrosinase activity and significantly downregulated the expressions of melanin synthesis-associated proteins, including microphthalmia-associated transcription factor (MITF), tyrosinase, dopachrome tautomerase (Dct), and tyrosinase-related protein-1 (TRP1). In conclusion, our findings indicated that activation of MEK/ERK and PI3K/Akt promoted by methyl gallate caused downregulation of MITF and triggered its downstream signaling pathway, thereby inhibiting the production of melanin. In summary, methyl gallate showed significant inhibitory activity against melanin formation, implying that it may be a potential ingredient for application in skin-whitening cosmetics.

Background: Although the link between non-steroidal anti-inflammatory drugs (NSAIDs) and tramadol and symptomatic hypoglycemia has been documented, there is a limited understanding of the associations of NSAIDs and tramadol with the risk of type 2 diabetes mellitus (T2DM). This study was established to evaluate the association between the clinical use of NSAIDs and the risk of T2DM. Patients and methods: A historical cohort study was conducted using the National Health Insurance Research Database in Taiwan dated from 2000 to 2013. Patients who received NSAIDs for at least 3 prescription orders and without co-treatment of tramadol in the exposure period (from 2000 to 2005) were considered as the exposed cohort (n = 3047). In comparison, patients who received tramadol for at least 3 prescription orders and without concomitant use of NSAIDs in the exposure period were considered as the comparison cohort (n = 9141). The primary outcome was the occurrence of T2DM. Multivariable hazard ratios (HRs) with 95% confidence intervals (CIs) derived from the Cox proportional hazard models were applied to determine the association between NSAIDs use and the risk of T2DM. Results: In the average follow-up period of 9.56 years, there were 159 newly diagnosed T2DM, with an incidence rate of 56.96 per 10,000 person years in the exposed cohort. Comparatively, there were 1737 incident T2DM cases, with an incidence rate of 161.23 per 10,000 person years in the comparison cohort. Compared to the comparison cohort, the NSAIDs cohort showed a significantly reduced risk of T2DM with an adjusted HR of 0.31 (95% CI, 0.26–0.36). Conclusions: Our cohort study provides longitudinal evidence that the use of NSAIDs was associated with a reduced risk of T2DM.

Background: Proton pump inhibitor (PPI) lansoprazole acts as a liver X receptor agonist, which plays a crucial role in the crosstalk of osteoblasts and osteoclasts in vitro and during bone turnover in vivo. However, epidemiological studies on the association between the use of lansoprazole and osteoporosis risk are limited. We aimed to determine the risk of developing osteoporosis in patients with lansoprazole use. Methods: A retrospective cohort study was conducted using the National Health Insurance Research Database of Taiwan dated from 2000 to 2013. The study includes 655 patients with lansoprazole use (the exposed cohort) and 2620 patients with other PPI use (the comparison cohort). The main outcome was the primary diagnosis of osteoporosis. The hazard ratios (HRs) and 95% confidence intervals (CIs) were used to assess the association between the use of lansoprazole and risk of osteoporosis. Results: Patients receiving lansoprazole treatment had a reduced risk of osteoporosis as compared with those undergoing other PPI therapy (adjusted HR, 0.56; 95% CI, 0.46–0.68). Moreover, this inverse association is evident in both sexes and in various age groups. Conclusions: This population-based cohort study demonstrated that lansoprazole use was associated with a reduced risk of osteoporosis. The clinical implications of the present study need further investigations.

Background/aim Urothelial carcinoma (UC) is the most common type of genitourinary cancer with high incidence and mortality rates in men. In this study, we used the BFTC-905 and T24 bladder cancer cell lines as in vitro models to investigate the pathways involved in flaccidoxide-induced apoptosis. Materials and methods We utilized MTT assays, colony assays, wound-healing assays and fluorescence with TUNEL to confirm the cytotoxicity of flaccidoxide in bladder cancer cell lines. Potential proliferative and apoptotic molecular mechanisms were evaluated by western blotting. Results The expression of anti-apoptotic proteins Bcl-2 and phosphorylated Bad (p-Bad) was attenuated with an increasing flaccidoxide concentration, while the expression of proapoptotic proteins Bax, Bad, cleaved caspase-3, cleaved caspase-9 and cleaved PARP-1 was found increased. Additionally, phosphorylation of phosphoinositide 3-kinases (PI3K), protein kinase B (AKT) and mammalian target of rapamycin (mTOR) in the PI3K/AKT/mTOR pathway was reduced, leading to a reduction in the phosphorylation of downstream 70-kDa ribosomal protein S6 kinase 1 (p70S6K), S6 ribosomal protein (S6) and eukaryotic translation initiation factor 4B (eIF4B). However, eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) protein phosphorylation was increased due to attenuation of the upstream phosphorylation of mTOR protein. Conclusion Flaccidoxide-induced apoptosis in BFTC-905 and T24 cells is mediated by mitochondrial dysfunction and down-regulation the PI3K/AKT/mTOR/p70S6K signaling pathway.

Background: Gout is the most common form of inflammatory arthritis in adults. Even though a link between gouty arthritis and type 2 diabetes mellitus (T2DM) has been reported, there is a limited understanding of the association between the anti-inflammatory agent colchicine and the risk of T2DM. This aim of this study was to assess the association between the use of colchicine and the risk of T2DM in an Asian cohort. Methods: A retrospective cohort study was conducted using the National Health Insurance Research Database (NHIRD) in Taiwan from 2000 to 2013. The study cohorts comprised 3841 gouty patients using colchicine (the exposed cohort) and 7682 gouty patients not using colchicine (the unexposed -cohort). The primary outcome was incident DM. The hazard ratios (HRs) and 95% confidence intervals (CIs) derived from a Cox proportional regression model were used to assess the association between colchicine use and the risk of diabetes. Results: The cumulative incidence of T2DM was significantly lower in the exposed cohort (18.8%) than in the unexposed cohort (25.0%). The risk of T2DM was significantly lower in colchicine users than in non-users (adjusted HR, 0.74; 95% CI, 0.36–0.87). The inverse relationship between colchicine use and diabetes risk remained consistent across sex and age groups. Conclusions: This cohort study provides longitudinal evidence that the use of colchicine is associated with a reduced risk of T2DM. This conclusion, however, needs to be interpreted cautiously given the lack of body mass index data in the NHIRD. Further studies are needed to determine the clinical implications of this study.

Background: Results of studies regarding the potential link between acid suppressant use and dementia risk are inconsistent. This study aimed to evaluate the association of cumulative exposure to histamine 2 receptor antagonists (H2RAs) and proton pump inhibitors (PPIs) with dementia risk in an Asian older cohort aged &ge, 65 years. Methods: Patients initiating H2RA (the H2RA user cohort, n = 21,449) or PPI (the PPI user cohort, n = 6584) and those without prescription for H2RA (the H2RA non-user cohort, n = 21,449) or PPI (the PPI non-user cohort, n = 6584) between 1 January 2000 and 31 December 2005 without a prior history of dementia were identified from Taiwan&rsquo, s National Health Insurance Research Database (NHIRD). The outcome of interest was all-cause dementia. Patients&rsquo, exposure to H2RAs or PPIs was followed-up from dates of initial prescription to the earliest outcome of incident dementia, death, or the end of 2013. Potential associations between acid suppressant use and dementia risk were analyzed using time-dependent Cox regression estimated hazard ratios (HRs) and 95% confidence intervals (CIs). Results: After mutual adjustment for H2RA and PPI use and other potential confounders, patients with H2RA use had significantly higher risk of developing dementia as compared to those not treated with H2RAs (adjusted HR, 1.84, 95% CI, 1.49&ndash, 2.20). Likewise, PPI users had significantly elevated risk of dementia compared to PPI non-users (adjusted HR, 1.42, 95% CI, 1.07&ndash, 1.84). Conclusions: Our results indicate that exposures to H2RAs and PPIs are associated with increased dementia risk.

Chondrosarcomas are well known for their resistance to chemotherapeutic agents, including cisplatin, which is commonly used in chondrosarcomas. Amphiregulin (AR), a ligand of epidermal growth factor receptor (EGFR), plays an important role in drug resistance. We therefore sought to determine the role of AR in cisplatin chemoresistance. We found that AR inhibits cisplatin-induced cell apoptosis and promotes ATP-binding cassette subfamily B member 1 (ABCB1) expression, while knockdown of ABCB1 by small interfering RNA (siRNA) reverses these effects. High phosphoinositide 3-kinase (PI3K), Akt and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) phosphorylation levels were observed in cisplatin-resistant cells. Pretreating chondrosarcoma cells with PI3K, Akt and NF-κB inhibitors or transfecting the cells with p85, Akt and p65 siRNAs potentiated cisplatin-induced cytotoxicity. In a mouse xenograft model, knockdown of AR expression in chondrosarcoma cells increased the cytotoxic effects of cisplatin and also decreased tumor volume and weight. These results indicate that AR upregulates ABCB1 expression through the PI3K/Akt/NF-κB signaling pathway and thus contributes to cisplatin resistance in chondrosarcoma.

Background Recent studies have raised concerns about the reduced efficacy of citalopram when used concurrently with proton pump inhibitors. The aim of this study was to evaluate the associations between clinical use of citalopram and omeprazole and the risk of sudden cardiac arrest ( SCA ) in an Asian population. Methods and Results A retrospective cohort study was conducted using the National Health Insurance Research Database of Taiwan dated from 2000 to 2013. The study cohorts comprised 3882 patients with citalopram use alone, 31 090 patients with omeprazole use alone, and 405 patients with concomitant use of citalopram and omeprazole (as the exposed cohort), and 141 508 patients received treatment with antidepressants without the risk of SCA and/or proton pump inhibitors other than omeprazole (as the comparison cohort). The primary outcome was the occurrence of SCA . The hazard ratios and 95% CI s derived from the time‐dependent Cox regression model were used to assess the association between the proposed drug treatments and risk of SCA . The adjusted hazard ratios of SCA was 1.32 (95% CI , 1.17–1.50) for citalopram use alone, 1.08 (95% CI , 0.98–1.20) for omeprazole use alone, and 2.23 (95% CI , 1.79–2.78) for concomitant use of citalopram and omeprazole. The cumulative incidence of SCA over the Kaplan‐Meier curves was more pronounced in patients with concomitant use of citalopram and omeprazole than those treated with citalopram alone and omeprazole alone. Conclusions This cohort study demonstrated use of citalopram and omeprazole either in isolation use or in concomitant use to be at increased risk for SCA .

Nobiletin (NOB) is a polymethoxylated flavonoid isolated from citrus fruit peel that has been shown to possess anti-tumor, antithrombotic, antifungal, anti-inflammatory and anti-atherosclerotic activities. The main purpose of this study was to explore the potential of using NOB to induce apoptosis in human bladder cancer cells and study the underlying mechanism. Using an MTT assay, agarose gel electrophoresis, a wound-healing assay, flow cytometry, and western blot analysis, this study investigated the signaling pathways involved in NOB-induced apoptosis in BFTC human bladder cancer cells. Our results showed that NOB at concentrations of 60, 80, and 100 &mu, M inhibited cell growth by 42%, 62%, and 80%, respectively. Cells treated with 60 &mu, M NOB demonstrated increased DNA fragmentation, and flow cytometry analysis confirmed that the treatment caused late apoptotic cell death. Western blot analysis showed that mitochondrial dysfunction occurred in NOB-treated BFTC cells, leading to cytochrome C release into cytosol, activation of pro-apoptotic proteins (caspase-3, caspase-9, Bad, and Bax), and inhibition of anti-apoptotic proteins (Mcl-1, Bcl-xl, and Bcl-2). NOB-induced apoptosis was also mediated by regulating endoplasmic reticulum stress via the PERK/elF2&alpha, /ATF4/CHOP pathway, and downregulating the PI3K/AKT/mTOR pathway. Our results suggested that the cytotoxic and apoptotic effects of NOB on bladder cancer cells are associated with endoplasmic reticulum stress and mitochondrial dysfunction.

In the previous study, the decolorization of azo dye was significantly enhanced by the light irradiation. The aim of this study is to investigate the effect of methyl orange decolorization under light-emitting diode (LED). The effect of media, pHs, and nitrogens on methyl orange decolorization by freely suspended cells are examined. The results showed that the optimal conditions for the methyl orange decolorization by theTsukamurellasp. J8025 are incubation at 30 °C and pH 8.0 in YEME medium under white-light LED.

Background: Ischemic stroke (IS) is a prevalent disease causing a body disability, the third leading cause of death in Taiwan. It shows that the level of intercellular adhesion molecular-1 (ICAM-1) in IS patients is higher than control subjects. Objective: This study is to investigate the possible association of ICAM-1 (G1548A) polymorphism in IS patients. Materials and Methods: A total of 646 subjects were enrolled in this study, including 312 IS patients, and 334 controls without a history of symptomatic IS. The ICAM-1 (G1548A) polymorphism was analyzed by polymerase chain reaction and restriction fragment length polymorphism. Clinical factors were also determined. Results: The frequencies of the ICAM-1 (G1548A) polymorphism for G/G, G/A, and A/A were 74.8%, 23.9%, and 0.3%, respectively, in healthy controls, and 62.8%, 32.1%, and 5.1%, respectively, in patients. The frequency of the ICAM-1 (G1548A) A allele (21.2% versus 13.2%, respectively; P = 0.007) and the carriers of the ICAM-1 (G1548A) A allele (37.2% versus 25.2%; P = 0.019, OR 1.76, 95% CI 1.1-2.83) are great in IS patients compared with healthy controls. There is a higher risk of IS associated with homozygosity for the ICAM-1 (G1548A) A allele (AA genotype) compared with the control population (5.1% vs. 0.3%, respectively, P = 0.04; OR 5.1, 95% CI 1.19-21.66). We also observed both hypertension and diabetes has shown a positive association with IS. Conclusions: The ICAM-1 (G1548A) polymorphism was associated with independent risk factor for the development of IS.

BACKGROUND AND OBJECTIVES: It has been proposed that glutamate decarboxylase 2 and the dopamine D2 receptor are involved in the brain reward cascade to increase carbohydrate craving and cause eating disorders. We investigated the association between the polymorphisms of the GAD2 and DRD2 genes and obesity with a higher body mass index (BMI) in Taiwanese patients. DESIGN AND SETTING: A retrospective, case-control study at Antai Tian-Sheng Memorial Hospital from 1 January to 31 December 2009. SUBJECTS AND METHODS: Of 300 subjects enrolled in the study, 132 were obese (BMI≥30 kg/m2) and 168 controls were not obese (BMI≤24 kg/m2). The polymorphisms of GAD2 (+61450 C/A), (+83987 T/A) and DRD2 (S311C) were characterized, respectively, by polymerase chain reaction-restriction fragment length polymorphism. The genotype and allele frequencies of the polymorphisms in this study were statistically analyzed. RESULTS: The genotype and allele frequencies of the GAD2 (+83987 T/A) and DRD2 (S311C) were significantly different between cases and controls (P=.001 for both). The frequencies of TT genotype and T allele of the GAD2 (+83987 T/A) as well as the frequencies of Ser/Cys genotype and Cys allele of DRD2 (S311C) were higher in cases compared to controls (P=.034 and .036 for both). CONCLUSIONS: The study demonstrated a statistically significant difference in the frequency of the GAD2 (+83987 T/A) and DRD2 (S311C) genes between cases and controls in Taiwanese subjects.

Background Noise-induced hearing loss (NIHL) is one of the common diseases in otology. In general, we assume that most people who are exposed to loud noise constantly, e.g., soldiers, will suffer from hearing loss. Hearing loss is related to the gene polymorphisms, with the Wolfram syndrome type 1 gene (T2500C), interleukin-4 receptor α chain (Q576R) and chloride channel-Kb (T481S) polymorphisms being most related to NIHL. Methods We analyzed the association between the polymorphisms and the risk of NIHL in 119 subjects who were exposed to the same loud gunfire. In the current study, 39 persons with hearing loss and 80 with normal hearing were recruited from military officers and soldiers that were exposed to gunfire. Results The results showed that the genetic variation of Wolfram syndrome type 1 gene was a more crucial factor than other genes in causing hearing loss. There was a significant difference ( p =0.027) in WFS1 (T2500C) polymorphism between NIHL subjects and controls. Conclusion In this study, we found that although loud noise could usually result in hearing damage, the clinical characteristics of hearing loss were irrelevant to gunfire noise. The gene polymorphisms provide predictors for us to evaluate the risk of NIHL prior to gunshot training.

In the previous study, the dye decolorization was investigated byTsukamurellasp. J8025 under the static condition at 30°C. The object of this study was to evaluate the influence of light irradiation with 15W low-pressure mercury lamp on dye decolorization. Three kinds of common culture medium Luria-Bertani (LB), Tryptic Soy Broth (TSB), and Yeast extract-Malt Extract (YEME) were used in this study. Strain J8025 was cultivated in different media added with methyl orange, and the rate of color removal was determined by measuring the absorbance at specific wavelengths. The experiments proved the decolorization efficiency after 48h under light irradiation in LB medium was up to 40%, that in TSB medium was up to 50%, and that in YEME medium was up to 68%, respectively. The decolorization process needed glucose as an energy source to support the bacterial growth and promote the decolorization rate. Due to the salt contained in the dye-wastewater, the effect of salt was investigated. The results showed nearly 98% color was removed after 48 h in the presence of 1% NaCl under light irradiation, but the decolorization was inhibited by high concentrations of salt. The results indicated a strain J8025 coupling with the light irradiation could be potentially used to improve the dye decolorization.

This study reports the statistical optimization of experiments, designed by Taguchi, to improve the wear-resistance of the tungsten carbide/cobalt-based coatings applied using high-velocity oxygen-fuel (HVOF) spraying processes. In this study, a group of needlelike textures appeared in the HVOF WC-Co coatings. The structures were evenly distributed and compacted closely, showing a more dense and homogenous configuration. The coating that formed with a large amount of pores, microcracks and unmolten particles was much looser, and the interface was locally prone to defects. The coatings were obviously not homogenous. Experimental results show that the wear-resistant coatings are highly dense, with an elastic lamellar structure. The tribological properties were significantly improved and the surface textures showed uniformity, in a molten droplet flattened formation. A homogenous, morphological structure, with local small aggregation, was more obvious after HVOF WC-Co coating. Accordingly, it is clear that the surface properties determined proved favorable to the mechanical properties of the WC/Co coatings. In addition, we compared the linear models with the quadric models, using RSM methods, in all the experiments. Statistically, the linear models produced an average error of 8.285％, while the quadric models generated an average error of 1.399％. It is clear that the RSM models successfully fitted the HVOF WC-Co coating process and the quadric models yielded more accurate predictions than the linear models, in most cases. The proposed procedure was applied to HVOF spraying of WC-CO coatings. The results demonstrated its feasibility and its effectiveness in improving the HVOF WC-Co coating.

A Rhodobacter sphaeroides WL-APD911 with the capability of producing the end products of neurosporene and ξ-carotene, rather than the normal end products of spheroidene, was isolated from hundreds of strains by using chemical mutagenesis. The strain WL-APD911 grew well in LB medium, and the colored carotenoids were produced within 3 days of incubation at 30°C. The Rs-M of colored carotenoid extracts obtained from strain WL-APD911 by using methanol extraction inhibited the NO production and iNOS expression by LPS-induced RAW 264.7 cells. This probability of reason for the Rs-M as the best source was to prompt the anti-inflammation and anti-oxidation in nutraceutical products. Finally, the carotenoid extracts (Rs-M) with neurosporene and ξ-carotene was called lycogene to be a newly biomaterial.

A Tsukamurella sp. J8025 with the capability of removing the synthetic dyes (methyl orange) was isolated from activated sludge. The strain J8025 grew well in aerobic and semi-aerobic condition, but the best decolorization was in semi-aerobic culture (static condition). For the optimal decolorization, the most suitable pH and temperature were pH 6.0 and 30°C. More than 80% of methyl orange was decolorized in the LB medium within 4 days of incubation. The decolorization rate could be strongly promoted to over 90% in LB media with glucose within 2 days of incubation. This probability of reason for the consumed glucose as the best energy source was to enhance the cell growth and decolorization activity.

Metastasis of cancer is the cause of the majority of cancer deaths. Active compound flaccidoxide-13-acetate, isolated from the soft coral Cladiella kashmani, has been found to exhibit anti-tumor activity. In this study, Boyden chamber analysis, Western blotting and gelatin zymography assays indicated that flaccidoxide-13-acetate exerted inhibitory effects on the migration and invasion of RT4 and T24 human bladder cancer cells. The results demonstrated that flaccidoxide-13-acetate, in a concentration-dependent manner, reduced the levels of matrix metalloproteinase-2 (MMP-2), MMP-9, urokinase-type plasminogen activator receptor (uPAR), focal adhesion kinase (FAK), phosphatidylinositide-3 kinases (PI3K), p-PI3K, AKT, p-AKT, mammalian target of rapamycin (mTOR), p-mTOR, Ras homolog gene family, member A (Rho A), Ras, mitogen-activated protein kinase kinase 7 (MKK7) and mitogen-activated protein kinase kinase kinase 3 (MEKK3), and increased the expressions of tissue inhibitor of metalloproteinase-1 (TIMP-1) and TIMP-2 in RT4 and T24 cells. This study revealed that flaccidoxide-13-acetate suppressed cell migration and invasion by reducing the expressions of MMP-2 and MMP-9, regulated by the FAK/PI3K/AKT/mTOR pathway. In conclusion, our study was the first to demonstrate that flaccidoxide-13-acetate could be a potent medical agent for use in controlling the migration and invasion of bladder cancer.

Previously, we showed that PMA activation of human monocyte-derived macrophages stimulates macropinocytosis (i.e., fluid-phase endocytosis) of LDL and transforms these macrophages into foam cells. The current study aimed to learn which PKC isoenzymes mediate cholesterol accumulation in PMA-activated human macrophages incubated with LDL. Cholesterol accumulation by PMA-activated macrophages incubated with LDL was nearly completely inhibited (85%) by the pan PKC inhibitors Go6850, Go6983, and RO 32-0432, but only was inhibited about 50% by the classical group PKC inhibitor, Go6976. This indicated that cholesterol accumulation was mediated by both a classical group and some other PKC isoenzyme. PKC beta was determined to be the classical group isoenzyme that mediated PMA-stimulated cholesterol accumulation. A pseudosubstrate myristoylated peptide inhibitor of PKC alpha and beta showed partial inhibition (congruent with 50%) of cholesterol accumulation. However, a small molecule inhibitor of PKC alpha, HBDDE, show minimal inhibition of cholesterol accumulation while a small molecule inhibitor of PKC beta, LY333513, could completely account for the inhibition of cholesterol accumulation by the classical group PKC isoenzyme. Thus, our findings show that beta and some other PKC isoenzyme, most likely delta, mediate cholesterol accumulation when macropinocytosis of LDL is stimulated in PMA-activated human monocyte-derived macrophages.

Regulation of intracellular protein stability by the ubiquitin-dependent proteasome system plays a crucial role in cell function. HO-1 (haem oxygenase) is a stress response protein, which confers cytoprotection against oxidative injury and provides a vital function in maintaining tissue homoeostasis. In the present study, we found a novel action of proteasome inhibitors MG132 and MG262 on HO-1 induction, and characterized the underlying mechanisms. MG132 (> or =0.1 microM) treatment resulted in a marked time- and concentration-dependent induction of the steady-state level of HO-1 mRNA in RAW264.7 macrophages, followed by a corresponding increase in HO-1 protein. Actinomycin D and cycloheximide inhibited MG132-responsive HO-1 protein expression, indicating a requirement for transcription and de novo protein synthesis. The involvement of signal pathways in MG132-induced HO-1 gene expression was examined using chemical inhibitors. Antioxidant N -acetylcysteine and SB203580, an antioxidant and inhibitor of p38 MAPK (mitogen-activated protein kinase), abolished MG132-inducible HO-1 expression. Furthermore, MG132 activated the p38 MAPK pathway. The half-life of HO-1 protein was prolonged by MG132, indicating that the upregulation of HO-1 by proteasome inhibitor is partially attributable to the inhibition of protein degradation. MG132 can ablate IkappaBalpha degradation and NF-kappaB (nuclear factor kappaB) activation induced by lipopolysaccharide, similar to the effect of another NF-kappaB inhibitor pyrrolidine dithiocarbamate. We found HO-1 upregulation by MG132 and pyrrolidine dithiocarbamate is unrelated to their inhibition of NF-kappaB, since leptomycin B, another NF-kappaB inhibitor, did not elicit similar induction of HO-1. Taken together, we found a novel effect of proteasome inhibitor on induction of HO-1 expression. This action is ascribed to the activation of the p38 MAPK pathway, but is not dependent on NF-kappaB inhibition.

Although capsaicin analogs might be a potential strategy to manipulate inflammation, the mechanism is still unclear. In this study, the effects and action mechanisms of vanilloid analogs on iNOS and COX-2 expression were investigated in RAW264.7 macrophages. Capsaicin and resiniferatoxin (RTX) can inhibit LPS- and IFN-γ-mediated NO production, and iNOS protein and mRNA expression with similar IC50 values of around 10 μM. Capsaicin also transcriptionally inhibited LPS- and PMA-induced COX-2 expression and PGE2 production. However, this effect exhibited a higher potency (IC50: 0.2 μM), and RTX failed to elicit such responses at 10 μM. Interestingly, we found that capsazepine, a competitive TRPV1 antagonist, did not prevent the inhibition elicited by capsaicin or RTX. Nevertheless, it mimicked vanilloids in inhibiting iNOS/NO and COX-2/PGE2 induction with an IC50 value of 3 μM. RT–PCR and immunoblotting analysis excluded the expression of TRPV1 in RAW264.7 macrophages. The DNA binding assay demonstrated the abilities of vanilloids to inhibit LPS-elicited NF-κB and AP-1 activation and IFN-γ-elicited STAT1 activation. The reporter assay of AP-1 activity also supported this action. The kinase assay indicated that ERK, JNK, and IKK activation by LPS were inhibited by vanilloids. In conclusion, vanilloids can modulate the expression of inflammatory iNOS and COX-2 genes in macrophages through interference with upstream signalling events of LPS and IFN-γ. These findings provide new insights into the potential benefits of the active ingredient in hot chilli peppers in inflammatory conditions. British Journal of Pharmacology (2003) 140, 1077–1087. doi:10.1038/sj.bjp.0705533

A segment of DNA containing the L-glutamate oxidase (gox) gene from Streptomyces platensis NTU3304 was cloned. The entire nucleotide sequence of the protein-coding portion consisting of 2130 bp (710 codons, including AUG and UGA) of the cloned DNA fragment was determined. The gox gene contained only one open reading frame (ORF) which coded for a 78-kDa polypeptide, the precursor of active extracellular Gox. Mature Gox is composed of three subunits, designated as α, β, and γ, with molecular masses of 39, 19, and 16 kDa, respectively. Analyses of the N-terminal amino acid sequences of the subunits revealed that the order of subunits in the precursor polypeptide encoded by the ORF, from N-terminus to C-terminus, is α–γ–β. The presence of the flavin adenine dinucleotide (FAD)-binding motif place Gox as a member of the flavoenzyme family. Furthermore, a negative effect of glucose on the biosynthesis of Gox was observed when it was used as carbon source.Key words: L-glutamate oxidase, gox gene, signal peptide, DNA sequence, flavoenzyme, pIJ702 vector.

A segment of DNA containing the L-glutamate oxidase (gox) gene from Streptomyces platensis NTU3304 was cloned. The entire nucleotide sequence of the protein-coding portion consisting of 2130 bp (710 codons, including AUG and UGA) of the cloned DNA fragment was determined. The gox gene contained only one open reading frame (ORF) which coded for a 78-kDa polypeptide, the precursor of active extracellular Gox. Mature Gox is composed of three subunits, designated as alpha, beta, and gamma, with molecular masses of 39, 19, and 16 kDa, respectively. Analyses of the N-terminal amino acid sequences of the subunits revealed that the order of subunits in the precursor polypeptide encoded by the ORF, from N-terminus to C-terminus, is alpha-gamma-beta. The presence of the flavin adenine dinucleotide (FAD)-binding motif place Gox as a member of the flavoenzyme family. Furthermore, a negative effect of glucose on the biosynthesis of Gox was observed when it was used as carbon source.

A 19-kb plasmid, pNI100, was isolated from Nocardia italica CCRC12359; its replicon was cloned and characterized as having a single open reading frame (ORF) of 1188 bp specifying 396 amino acids (aa). Analyses of the deduced aa sequence of the Rep protein indicated that characteristics of three consensus sequences and a P-loop-like motif in the Rep protein of plasmid pSG5, a conjugative plasmid involving a rolling-circle replication mechanism, were conserved in those of plasmid pNI100. Phenotypically, a pock structure was produced in the regenerated mycelium by introducing pNI100 DNA into the Streptomyces lividans protoplast. This result strongly suggests that pNI100 is a conjugative plasmid and probably replicates by a rolling-circle replication mechanism. By using the replicon of pNI100, a bifunctional plasmid pNI105 that could replicate in both Escherichia coli and S. lividans was constructed and found to be a useful cloning shuttle vector.

The aim of this study is to examine the influence of Light-emitting diode (LED) on the decolorization of azo dyes. A Tsukamurella sp. J8025 with the best capability of removing the methyl orange was cultivated in static condition. Yeast extract-Malt Extract (YEME) was used in this study. White LEDs was used as light source in this study. The decolorization rate was determined by measuring the absorbance at 485 nm wavelength. The experimental results proved the decolorization efficiency by using J8025 strain cultivated in YEME medium under white and blue LED at 30°C was up from 1.3 to 1.5 fold compared to the control without lighting. In addition, the decolorization rate could be promoted in YMEM medium with 1% of NaCl concentration. The possible reason for the ion effect caused from NaCl could reduce the solubility of azo dyes and increase the chance of strain J8025 contacting the dyes.

Background and Objectives: Osteoporosis is a common disease of the elderly, in which genetic and clinical factors contribute to the disease phenotype. Since the production of interleukin-1 (IL-1) has been implicated in the bone mass and skeletal disorders, we investigated whether IL-1 system gene polymorphisms are associated with the pathogenesis of osteoporosis in postmenopausal Taiwanese women. Methods: Osteoporosis is diagnosed by dual-energy x-ray absorptiometry, which measures bone mineral density (BMD) at multiple skeletal sites. We studied the IL-1α (-889C/T), IL-1β (-511C/T) and the 86 base pair variable number tandem repeat (VNTR) in intron 2 of the IL-1 receptor antagonist (IL-1ra) gene in 117 postmenopausal women with osteoporosis and 135 control subjects without a history of symptomatic osteoporosis. These gene polymorphisms were analyzed by polymerase chain reaction and restriction fragment length polymerase. Blood sugar and other risk factors were also determined. Results: The frequencies of IL-1β (-511C/T) genotypes (P=.022, odds ratio=1.972) and alleles (P=.02, odds ratio=2.909) showed a statistically significant difference between the two groups. However, we did not find any statistically significant difference in IL-1β and IL-1ra polymorphisms (P>.05). We also observed a positive relationship between osteoporosis and cholesterol and a weak inverse relationship between blood sugar and osteoporosis in postmenopausal women. Conclusions: These experimental results suggest that the pathogenesis of osteoporosis is associated with IL-1β (-511C/T) polymorphism in postmenopausal women. This polymorphism is an independent risk factor for osteoporosis.

In the present study, experiments were performed to explore the action of quercetin, the most widely distributed flavonoids, and its major metabolite, quercetin-3'-sulfate, on lipopolysaccharide (LPS)- and interferon-gamma (IFN-gamma)-induced nitric oxide (NO) production in BV-2 microglia. Quercetin could suppress LPS- and IFN-gamma-induced NO production and inducible nitric oxide synthase (iNOS) gene transcription, while quercetin-3'-sulfate had no effect. LPS-induced IkappaB kinase (IKK), nuclear factor-kappaB (NF-kappaB) and activating protein-1 (AP-1) activation, and IFN-gamma-induced NF-kappaB, signal transducer and activator of transcription-1 (STAT1) and interferon regulatory factor-1 (IRF-1) activation were reduced by quercetin. Moreover quercetin was able to induce heme oxygenase-1 expression. To address the involvement of heme oxygenase-1 induction in iNOS inhibition, heme oxygenase-1 antisense oligodeoxynucleotide was used. Quercetin-mediated inhibition of NO production and iNOS protein expression were partially reversed by heme oxygenase-1 antisense oligodeoxynucleotide, but was mimicked by hemin, a heme oxygenase-1 inducer. The involvement of signal pathways in quercetin-induced heme oxygenase-1 gene expression was associated with tyrosine kinase and mitogen-activated protein kinases activation. All these results suggest quercetin should provide therapeutic benefits for suppression of inflammatory-related neuronal injury in neurodegenerative diseases.

Except functioning as lipid-lowering agents, HMG-CoA inhibitors, statins, are good tools to clarify the signaling role of small G proteins. In this study, we found in murine RAW264.7 macrophages, statins within 1-30 microM stimulated COX-2 gene transcription and PGE(2) formation, displaying potencies as lovastatin > fluvastatin > atorvastatin >> pravastatin. Transfection experiments with COX-2 promoter construct showed the necessity of C/EBPbeta and CRE promoter sites, but not NF-kappaB promoter site. Effects of statins on the activation of COX-2 promoter, induction of COX-2 protein, and PGE(2) production were all prevented by mevalonate and prenylated metabolites, FPP and GGPP. Consistent with the effect of statins, manumycin A, farnesyltransferase inhibitor, and geranylgeranyltransferase inhibitor increased PGE(2) production and COX-2 induction. Likewise, toxin B, an inhibitor of Rho family members, caused a prominent COX-2 induction. Results also indicated that tyrosine kinase, ERK, and p38 MAPK play essential roles in statin action. Taken together, these results not only demonstrate a unique action of statins in the upregulation of COX-2 expression in macrophages, but also suggest a negative role controlled by small G proteins in COX-2 gene regulation. Removal of this negative control by impairing G protein prenylation with statins leads to MAPKs activation and promotes COX-2 gene expression through the activation at CRE and C/EBPbeta sites.

Calcium/calmodulin-dependent protein kinase kinase (CaMKK) and Akt are two multifunctional kinases involved in many cellular responses. Although Akt and Ca(2+) signals have been implicated in NF-kappaB activation in response to certain stimuli, these results are still controversial, and the mechanism(s) involved remains unknown. In this study, we show the roles that CaMKK and Akt play in regulating interleukin-1beta (IL-1beta)-induced NF-kappaB signaling. In human embryonic kidney 293 cells, IL-1beta induces IkappaB kinase beta (IKKbeta) activation, IkappaBalpha degradation, NF-kappaB transactivation, and weak Akt activation. A CaMKK inhibitor (KN-93) and phosphatidylinositol 3-kinase inhibitors (wortmannin and LY294002) do not inhibit IL-1beta-induced NF-kappaB activation. However, IL-1beta-induced NF-kappaB activity is attenuated by increased intracellular calcium in response to ionomycin, UTP, or thapsigargin or by overexpression of CaMKKc and/or Akt. Ionomycin and CaMKKc overexpression increases Akt phosphorylation on Thr(308) and enzyme activity. Under these conditions or upon overexpression of wild type Akt, IL-1beta-induced IKKbeta activity is diminished. Furthermore, a dominant negative mutant of Akt abolishes IKKbeta inhibition by CaMKKc and ionomycin, suggesting that Akt acts as a mediator of CaMKK signaling to inhibit IL-1beta-induced IKK activity at an upstream target site. We have also identified a novel interaction between CaMKK-stimulated Akt and interleukin-1 receptor-associated kinase 1 (IRAK1), which plays a key role in IL-1beta-induced NF-kappaB activation. CaMKKc and Akt overexpression decreases IRAK1-mediated NF-kappaB activity and its association with MyD88 in response to IL-1beta stimulation. Furthermore, CaMKKc and Akt overexpression increases IRAK1 phosphorylation at Thr(100), and point mutation of this site abrogates the inhibitory effect of Akt on IRAK1-mediated NF-kappaB activation. Taken together, these results indicate a novel regulatory mechanism for IL-1beta signaling and suggest that CaMKK-dependent Akt activation inhibits IL-1beta-induced NF-kappaB activation through interference with the coupling of IRAK1 to MyD88.

Objective: Broussonetia papyrifera is used as a traditional medicine to treat few diseases. However, the antiinflammatory effect of B. papyrifera stem bark has not been evaluated. The aim of this study is to investigate the effects of n-hexane fraction from methanol extract of B. papyrifera stem bark on lipopolysaccharide (LPS)-stimulated inflammation using RAW 264.7 cells. Materials and Methods: Methanol extract was obtained from B. papyrifera stem bark and its sequential fractions (hexane, dichloromathane, ethyl acetate, butanol, and aqueous) were obtained by extraction in solvents with increasing polarity and were examined in RAW 264.7 cells. Results: The secretion profiles of pro-inflammatory parameters, including nitric oxide (NO), tumor necrosis factor-α (TNF-α), and interleukin-1β (IL-1β) were found to be significantly reduced in 10-80 μg/ml dose ranges of n-hexane fraction (BP-H) from methanol extract of B. papyrifera stem bark. The expressions of inducible NO synthase (iNOS) was also significantly inhibited by BP-H. Reverse transcription-polymerase chain reaction (RT-PCR) analysis showed that BP-H treatment decreased LPS-induced iNOS mRNA expression in RAW 264.7 cells. Conclusion: The results suggest that the B. papyrifera stem bark has anti-inflammatory activity which inhibits the NO production and proinflammatory cytokines in RAW 264.7 cells. B. papyrifera stem bark might act as a potential therapeutic agent for inflammatory diseases.

A concentric-cylinder airlift reactor, in which the annulus is a packed bed of glass fibers, has been developed in order to facilitate the scaleup and enhance the volumetric productivity of anchorage-dependent animal cell cultures. In this bio-reactor, oxygen-containing gas is sparged through the inner draft tube, causing bubble-free medium to flow through the fiber bed in the outer cylinder and providing both oxygenation and convective nutrient transfer to the cells. Several other desirable features for reactor operation are also provided by this design. Cell cultivations in this bioreactor have been successfully carried out and provide data for the feasibility of the large-scale cell cultivation.