The nuclear translocation of STAT1 and STAT2 is a key process in IFN-α transduction signaling. We and others have shown that STAT1 translocation is impaired in Pol-expressing and HBV-infected hepatocytes.6, learn more 7 We further examined whether Pol interferes with IFN-α–induced STAT2 nuclear accumulation. In the absence of IFN-α, STAT2 was predominantly localized in the cytoplasm, whereas STAT1 was found in both the cytoplasm and nucleus; upon IFN-α stimulation, strong nuclear accumulation of both STAT1 and STAT2 was

observed, but only in cells without Pol expression (Fig 4A). Furthermore, the protein levels of STAT1/2 in the cytoplasmic and nuclear fractions of Dox-treated or Dox-free HepAD38 cells were determined via immunoblotting. As shown in Fig. 4B, the accumulation of STAT1/2 in the nucleus following IFN-α treatment was significantly detained in Dox-free (Pol-expressing) cells. Impaired nuclear translocation

SCH772984 chemical structure of STAT1/2 was also observed in HepG2.215 cells compared with HepG2 cells (Supporting Fig. 6). Importin-α5 (also known as karyopherin α1), a nuclear localization signal receptor, has been shown to specifically interact with the STAT1-STAT2 heterodimer and to be responsible for the nuclear transport of the complex.17, 18 We thus investigated whether Pol interferes with the interaction between importin-α5 and activated STATs. As shown in Fig. 5A, STAT1 and STAT2 were clearly detected in the importin-α5 immunoprecipitation find more complex when IFN-α was added, however, the STATs decreased in a dose-dependent manner, with increased expression of Pol. A similar inhibition was observed using the HepAD38 model (Supporting Fig. 5C). Moreover, impaired colocalization of STAT2 and importin-α5 was observed in HepG2.215 cells compared with that in HepG2 cells by immunofluorescence

(Fig. 5B). To investigate whether Pol directly interacts with importin-α5, GST pull-down assays were conducted, and Flag-Pol was pulled down by GST–importin-α5 (Fig. 5C). Furthermore, colocalization of Pol and importin-α5 in the cytoplasm was detected (Fig. 6D). The C-terminal arm repeats 8 and 9 of importin-α5 have been reported to form a unique binding site for activated STAT1-STAT2.19 Thus, we aimed to determine whether Pol binds to this region of importin-α5. Hemagglutinin-tagged full-length importin-α5 and several truncated constructs were transfected along with Flag-Pol, and the co-IP results showed that Pol was able to coprecipitate with all the truncations except importin-α5-1-406 (Fig. 5D), implying that Pol binds to importin-α5 through a region (407-504) that is also required for the importin-α5-STAT1/2 interaction.

Within these patients, 13 lesions in 12 patients who developed local tumor progression in the follow-up period of at least 8 months were retrospectively reviewed. Imaging obtained before and after RFA was used for creating fused images on a workstation. Ablative margins were assessed using only axial images, and with fused images. The ablative margin was assessed as sufficient in all 13 lesions using side-by-side axial images; however, all lesions

were assessed as insufficient with fused imaging evaluation. The reason for the discrepancy of the assessment results were differences in the respiratory dislocation of the liver in the pre- and Obeticholic Acid order post-RFA images in eight lesions (61.5%), and rotational displacement of the liver and the torso in five (38.5%). The site of local tumor recurrence relative to the HCC lesion was craniocaudal in 12 lesions, dorsoventral in seven and lateral in seven. In all lesions, the site of local tumor recurrence was congruent with the area of the thinnest ablative margin. Selleck MS275 Assessment of ablative margin with fused imaging revealed insufficiency of ablation previously evaluated as sufficient with conventional axial imaging. Fused imaging evaluation has proved to be an accurate and useful tool for the assessment of RFA margins. “
“Mice deficient in small heterodimer partner (SHP) are protected from diet-induced hepatic steatosis resulting

from increased fatty acid oxidation and decreased lipogenesis. The decreased lipogenesis appears to be a direct consequence of very low expression of peroxisome proliferator-activated receptor gamma 2 (PPAR-γ2), a potent lipogenic transcription factor, in the SHP−/− liver. The current

study focused on the identification of a SHP-dependent regulatory cascade that controls PPAR-γ2 gene expression, thereby regulating hepatic fat accumulation. Illumina BeadChip array (Illumina, Inc., San Diego, CA) and real-time polymerase chain reaction were used to identify genes responsible for the linkage between SHP and PPAR-γ2 using selleck screening library hepatic RNAs isolated from SHP−/− and SHP-overexpressing mice. The initial efforts identify that hairy and enhancer of split 6 (Hes6), a novel transcriptional repressor, is an important mediator of the regulation of PPAR-γ2 transcription by SHP. The Hes6 promoter is specifically activated by the retinoic acid receptor (RAR) in response to its natural agonist ligand, all-trans retinoic acid (atRA), and is repressed by SHP. Hes6 subsequently represses hepatocyte nuclear factor 4 alpha (HNF-4α)-activated PPAR-γ2 gene expression by direct inhibition of HNF-4α transcriptional activity. Furthermore, we provide evidences that atRA treatment or adenovirus-mediated RAR-α overexpression significantly reduced hepatic fat accumulation in obese mouse models, as observed in earlier studies, and the beneficial effect is achieved by the proposed transcriptional cascade.

Within these patients, 13 lesions in 12 patients who developed local tumor progression in the follow-up period of at least 8 months were retrospectively reviewed. Imaging obtained before and after RFA was used for creating fused images on a workstation. Ablative margins were assessed using only axial images, and with fused images. The ablative margin was assessed as sufficient in all 13 lesions using side-by-side axial images; however, all lesions

were assessed as insufficient with fused imaging evaluation. The reason for the discrepancy of the assessment results were differences in the respiratory dislocation of the liver in the pre- and ATM/ATR mutation post-RFA images in eight lesions (61.5%), and rotational displacement of the liver and the torso in five (38.5%). The site of local tumor recurrence relative to the HCC lesion was craniocaudal in 12 lesions, dorsoventral in seven and lateral in seven. In all lesions, the site of local tumor recurrence was congruent with the area of the thinnest ablative margin. selleck chemical Assessment of ablative margin with fused imaging revealed insufficiency of ablation previously evaluated as sufficient with conventional axial imaging. Fused imaging evaluation has proved to be an accurate and useful tool for the assessment of RFA margins. “
“Mice deficient in small heterodimer partner (SHP) are protected from diet-induced hepatic steatosis resulting

from increased fatty acid oxidation and decreased lipogenesis. The decreased lipogenesis appears to be a direct consequence of very low expression of peroxisome proliferator-activated receptor gamma 2 (PPAR-γ2), a potent lipogenic transcription factor, in the SHP−/− liver. The current

study focused on the identification of a SHP-dependent regulatory cascade that controls PPAR-γ2 gene expression, thereby regulating hepatic fat accumulation. Illumina BeadChip array (Illumina, Inc., San Diego, CA) and real-time polymerase chain reaction were used to identify genes responsible for the linkage between SHP and PPAR-γ2 using selleck inhibitor hepatic RNAs isolated from SHP−/− and SHP-overexpressing mice. The initial efforts identify that hairy and enhancer of split 6 (Hes6), a novel transcriptional repressor, is an important mediator of the regulation of PPAR-γ2 transcription by SHP. The Hes6 promoter is specifically activated by the retinoic acid receptor (RAR) in response to its natural agonist ligand, all-trans retinoic acid (atRA), and is repressed by SHP. Hes6 subsequently represses hepatocyte nuclear factor 4 alpha (HNF-4α)-activated PPAR-γ2 gene expression by direct inhibition of HNF-4α transcriptional activity. Furthermore, we provide evidences that atRA treatment or adenovirus-mediated RAR-α overexpression significantly reduced hepatic fat accumulation in obese mouse models, as observed in earlier studies, and the beneficial effect is achieved by the proposed transcriptional cascade.

We further investigated whether the MESIAH score can further classify survival of patients within each stratum of the BCLC staging system (Fig. 4). Whereas patients in BCLC stage 0 or A did well overall, there was a large degree of variability in survival of patients with BCLC stage B to D. In BCLC stage B, patients in the lowest quartile of the MESIAH score had 77% 3-year survival compared with those in highest quartile with 17% survival. The gap was even wider in BCLC stage C patients, in whom the 3-year survival in the lowest

and highest quartiles was 49% and 0%, respectively. Sensitivity analyses were performed to test the robustness of the MESIAH score. First we examined the effect of censoring patients at the time of liver transplantation (n = 133), as the Sotrastaurin cost procedure may fundamentally affect the natural history of HCC. The result, however, was that censoring liver transplantation did not affect the overall performance of the model. Dasatinib order The c-statistic in the derivation cohort was 0.79 and validation 0.81, which were again better than other staging systems (Supporting Table). Second, we repeated

the validation analysis excluding nonviral hepatitis patients in the validation cohort. Again, the result did not change materially (data not shown). In general, patient characteristics that determine the prognosis in HCC tend to be more complex than those for other solid tumors, as the vast majority of patients with HCC have underlying liver cirrhosis and the degree of hepatic dysfunction, in addition to the extent of the tumor, affects prognosis. This analysis demonstrates that the extent of the tumor as represented by variables such as the number and size of the lesions, vascular invasion, and extrahepatic metastasis and the underlying liver function, measured by MELD, are important independent predictors of survival in HCC patients.

Although this is not learn more the first observation that MELD may be useful in prognosticating patients with HCC,19 one of the advantages of the MESIAH score is that it only includes objective, reproducible variables. The value of an objectively quantifiable measure of disease has been appreciated with MELD, which has been rapidly become a common language among physicians globally. Similarly, we believe that the MESIAH score could be applied in retrospective studies or in epidemiologic research where nuanced details of clinical information are unavailable. We are reassured of the validity of the MESIAH score model, as it performed well in our cross-validation as well as in the independent dataset of our validation cohort. The c-statistic in the validation cohort was higher than that in the derivation cohort, indicating that it was easier for the score to separate HCC patients according to their prognosis in the former. This is likely attributed to the wider range of the score (and thus survival) with fewer patients receiving treatment that potentially alters the natural history in that cohort.

022, respectively, Fisher’s exact test) (Fig. 7). There was, however, no association between SP1 and MMP2 expression using immunohistochemistry (P = 0.740). No association between the clinicopathologic features and SP1 and

MMP2 overexpression was found. Despite the fact that PTEN has been extensively studied and is implicated in cell migration,15-19 its underlying molecular mechanisms in HCC progression and metastasis have not been clearly learn more elucidated. Therefore, it is of strategic importance to characterize the functional consequence of PTEN underexpression and the deregulated downstream signaling. In line with our clinical findings, the metastatic cell line H2M had a lower PTEN protein expression compared with its primary HCC counterpart cell line H2P.10 To study the functional consequences of PTEN loss in HCC, two HCC cell lines, SMMC-7721 and BEL-7402, with relatively higher endogenous PTEN levels were used to establish the stable PTEN-knockdown clones. Selleckchem Alvelestat Because more than one stable knockdown clone was used, this likely eliminated clonal effect in the assays. We documented that knockdown of PTEN significantly promoted cell migration and invasion

in vitro, suggesting its possible role in HCC metastasis. Similar results were obtained with both cell lines, which suggests that the enhanced cell migration and invasion associated with loss of PTEN expression was not cell line–specific. In addition, our PTEN-deficient MEFs also exhibited increased cell migration and invasion compared with wild-type

MEFs. This indicates that the association of PTEN loss with metastasis is not cell type–specific. Overall, our in vitro results were consistent with our clinical observation that PTEN underexpression was significantly associated with more frequent tumor microsatellite formation in human HCCs. Tumor microsatellite formation in human HCCs is one of the histologic features of tumor metastasis. Furthermore, we observed that the PTEN−/− MEFs also possessed enhanced check details proliferation rate (Supporting Fig. 2), and this was consistent with our finding of p-AKT activation, thereby triggering the prosurvival pathways. The enhanced cell proliferation is in agreement with our finding of an association of PTEN underexpression with increased tumor size in human HCCs. To eliminate the additive effect or cell proliferation in contribution to the enhanced cell migration and invasion observed, mitomycin C, a drug that blocks cell proliferation, was added in these assays. To move across the Matrigel layer in the cell invasion assay, cells need to secrete certain enzymes to degrade the extracellular matrix. Indeed, the Matrigel coated onto the invasion chamber in the cell invasion assay comprise three main substances: 56% laminin, 31% collagen IV, and 8% entactin, all by weight. MMP2 and MMP9 gelatinases primarily act to destroy the major constituent collagen IV.

However, the bioavailability of cyclosporine varies considerably depending on patient population buy Roxadustat (ranging from <10% in liver transplant patients to 89% in some kidney transplant patients).18 Therefore, the effect of telaprevir on cyclosporine concentrations in liver transplant patients may differ from that observed in this healthy volunteer study, and close monitoring of cyclosporine concentrations to guide individual dose adaptations would be necessary

during coadministration. The decrease in hepatic clearance and increase in t½ of both cyclosporine and tacrolimus upon telaprevir coadministration suggests that systemic clearance of these immunosuppressants was also reduced by telaprevir. The effect of telaprevir on hepatic transporters that could have contributed to lower clearance or enhanced absorption is unknown. Notably, in this study the effect of steady-state telaprevir on the PK of cyclosporine or tacrolimus was evaluated only at single doses of these immunosuppressants.

Because the elimination half-lives increased significantly for both cyclosporine and tacrolimus when telaprevir was coadministered, without proper adjustment of dose and dosing interval of these immunosuppressants, further increases in blood exposure may occur when multiple doses of these drugs are coadministered with telaprevir. However, studies of telaprevir with multiple doses of cyclosporine and tacrolimus have not been performed. The effects of

telaprevir on cyclosporine and tacrolimus PF-02341066 in vitro exposure were similar to that reported for human immunodeficiency virus (HIV) protease inhibitors known to be potent CYP3A inhibitors, where significant reductions in dose and/or dosing interval of immunosuppressants were needed to achieve the desired range of trough concentrations, based on frequent monitoring of trough concentrations of the immunosuppressants.25 For example, addition of lopinavir/ritonavir (n = 7 patients) reduced tacrolimus learn more dose by 99% to maintain tacrolimus concentrations within the therapeutic range.26 Similarly, during coadministration of Highly Active Antiretroviral Therapy (HAART) regimens with ritonavir-boosted HIV protease inhibitors, daily cyclosporine doses were reduced by 80%-95% to maintain cyclosporine exposure at pre-HAART levels. Because of the flat absorption/elimination profiles of cyclosporine during combination with ritonavir-boosted HAART therapy, cyclosporine exposure could be reliably monitored long-term by measuring cyclosporine trough concentrations.27 Treatment of posttransplant patients coinfected with HIV/HCV with antiretrovirals and telaprevir could be even more challenging, depending on the drugs involved. Telaprevir levels are not significantly affected by ritonavir28; however, whether the net effect of antiretroviral drugs on cyclosporine and tacrolimus PK would be similar or different is hard to predict, as these drugs may have their own effects.

Conclusion: Temporary placement of a FSCEMS in the PD for aiding extraction of large PD stones is a safe technique that facilitates the removal of large stones. Key Word(s): 1. pancreas; 2. metal stent; 3. stones; Presenting Author: ENQIANG LINGHU Additional Authors: YOU ZHANG, LIHUA PENG, XIAOLIN SHI, YONGWEI ZHAO, QIYANG HUANG, CHEN DING, XIAOYU QIU Corresponding Author: ENQIANG LINGHU Affiliations: Department of Gastroenterology and Hepatology, the Chinese PLA General Hospital;

Department of Gastroenterology and Hepatology, the PLA General Hospital; Department of Gastroenterology and Hepatology, Chinese PLA General Hospital Objective: Peroral endoscopic myotomy (POEM) in combination with balloon shaping is a new treatment for selleck products achalasia. The aim of our study was to determine the efficacy and safety of POEM in combination with balloon shaping in the treatment of patients with achalasia Methods: Symptom relief rate, changes in body weight, changes in esophageal sphincter this website residual pressure (ESRP) and complication rate of 15 patients with achalasia before and after treatment were retrospectively analyzed.

The follow-up time is 3 months. Results: The POEM in combination with balloon shaping significantly reduced the symptom score in all cases (from mean 7.8 to 0.53, P = 0.000) and the symptom relief rate was 100%. The post-treatment average body weight of 15 patients was significantly higher than that of before (62.9 VS 59.6, P = 0.0003). The treatment significantly improved the lower esophageal sphincter residual pressure (LESRP) (from mean 19.1 mmHg to 12.3 mmHg, P = 0.0059) and upper esophageal sphincter residual pressure (UESRP) (from mean 16.1 mmHg to 5.1 mmHg, P = 0.0365) in 5 cases who had checked the esophageal motility in three months after operation. There was one case (1/15, 6.7%) of pneumoperitoneum

during operation and one case (1/15, 6.7%) of reflux esophagitis in 3 months after treatment Conclusion: The POEM in combination with balloon shaping can significantly improved the symptoms of patients with achalasia in the short term and is safe for the treatment of patients with achalasia. Further studies are needed to confirm its long-term efficacy in patients with achalasia. Key Word(s): 1. POEM; 2. balloon shaping; 3. achalasia; Presenting Author: KWUNG selleckchem JUN PARK Additional Authors: YOUNG SOO PARK, CHEOL MIN SHIN, SANG HOON JEON, HEE JIN KIM, NAYOUNG KIM, DONG HO LEE Corresponding Author: KWUNG JUN PARK Affiliations: Department of Internal Medicine, Seoul National University Bundang Hospital; Department of Department of Thoracic and Cardiovascular Surgery Objective: Over the past 20 years photodynamic therapy (PDT) has become a viable treatment method for early and developing stages of esophageal cancer. Our study examined the outcome of esophageal squamous cell carcinoma by using porfimer sodium (Photofirn, photogem, Photodin), radachlorin and aminolevulinic acid (ALA) -mediated PDT.

1A and 1B. In these figures, it is demonstrated that many C282Y homozygotes have normal ALT levels, but also that patients with an elevated ALT level are unlikely to be C282Y homozygotes. The correlation between ALT and ferritin was stronger in C282Y homozygotes than in nonhomozygotes, which is consistent with an inflammatory cause of the hyperferritinemia in nonhomozygotes. Buparlisib The proportion of male C282Y homozygotes with ALT and AST levels <40 IU/L was 71% and 87%, respectively. The proportion of female C282Y homozygotes with ALT and AST levels <40 IU/L was 87%

and 95%, respectively. The decreasing probability of being a C282Y homozygote across groups in men and women with increasing ALT levels is shown in Fig. 2. Similar results were determined for AST. P values for chi-square tests for trends in proportions for ALT were 0.036 for men and 0.00017 for women. Mantel-Haenszel chi-square adjusted for gender was <0.0001. An unanticipated observation was that the probability of being a C282Y homozygote decreased as serum ALT and AST levels increased. The results of the subgroup analysis limited to Caucasians were similar. It is widely believed that the probability of diagnosing many liver diseases increases as serum transaminases increase. In the present study of subjects with

hyperferritinemia, the probability of being a C282Y homozygote decreased with increasing ALT and AST levels. This probably occurs

because the deposition of excessive iron alone in hepatocytes of persons with hemochromatosis is not inflammatory. “Silent” hepatic fibrosis Selinexor occurs in some subjects with hemochromatosis and normal serum transaminases.6, 7 On the other hand, some patients with hemochromatosis and HFE C282Y homozygosity have both hepatic iron overload and an inflammatory liver condition. For example, approximately 15% of C282Y homozygotes diagnosed in medical care selleck products have severe hepatic steatosis proven by liver biopsy. These patients had higher median serum ALT and ferritin levels than C282Y homozygotes without hepatic steatosis or other inflammatory liver disorder.8 In contrast, patients referred for evaluation of elevated serum ferritin levels usually have hyperferritinemia resulting from inflammatory liver disease, rather than iron overload resulting from HFE hemochromatosis.9 In prospective analyses of subjects with chronic elevation of serum transaminases, hepatic steatosis associated with or without excessive ethanol consumption was the predominant cause of elevated serum transaminases.10-13 Hemochromatosis was rare in these case series.9 In the present study, there was a potential bias wherein HEIRS Study non-C282Y homozygous participants were deliberately selected for postscreening clinical examinations because they had elevated serum transferrin saturation and ferritin measures.

31)8 were synthesized in the

Medicinal Chemistry Laboratory of the Institute of Medicinal Biotechnology Chinese Academy of Medical Sciences, with a purity over 99.0%. The compound structure was confirmed with 1H-NMR and MS spectra. Interferon-α-2b (Intron A) was from Schering Plough (Brinny) (Kenilworth, NJ). BILN2061, a known NS3-4A protease inhibitor, was provided by Shanghai Lechen International Trading (Shanghai, China). Plasmid pcDNA3.1-Vif coding for HIV-1 full-length Vif was created by insertion of Vif (amplified selleck screening library by polymerase chain reaction [PCR] from HIV-1 plasmid SVC21.BH10) into pcDNA3.1; the plasmids hA2, hA3B, hA3C, hA3F, and hA3G that express wildtype forms of hA2, hA3B, hA3C, hA3F, and hA3G, respectively, possess a fused HA tag at the C-terminus. The above-mentioned plasmids were gifts from Dr. Shan Cen at the Lady Davis Institute for Medical Research and McGill University AIDS Centre. The plasmid pFL-J6/JFH/JC1 containing the full-length chimeric

HCV cDNA was kindly provided by Vertex Pharmaceuticals (Boston, MA). Production of infectious HCV in hepatocytes was done as described.14 The plasmid pFL-J6/JFH/JC1 was restricted with XbaI and treated with Mung Bean nuclease (New England Biolabs) to generate the according HCV cDNA with T7 promotor. The cDNAs were purified and used as templates for RNA synthesis. HCV RNA was synthesized selleck compound in vitro using a MEGAscript T7 kit (Ambion). The synthesized RNA was treated

with DNase I (New England Biolabs) and purified with Wizard check details SV Gel and PCR Clean-Up System (Promega). The synthesized HCV RNA was used to transfect naïve Huh7.5 cells with the addition of Lipofectamine 2000 (Invitrogen). The culture medium was collected and cleaned with centrifugation at 3,000 rpm for 10 minutes. The supernatants were stored at −70°C as HCV viral stock and quantified with the Diagnostic Kit for Quantification of Hepatitis C Virus RNA (Shanghai Kehua Bio-Engineering). Huh7.5 cells 24 hours after HCV infection with viral stock (45 IU per cell) were transfected with different concentrations of APOBEC- or Vif-containing plasmids in the FuGENE HD Transfection Reagent (Roche), with pcDNA3.1 as plasmid control. Then, 72 hours later the culture medium was removed and total intracellular proteins were extracted using CytoBuster Protein Extraction Reagent (Novagen) with 1 mM protease inhibitor cocktail (Roche Applied Science). HCV Core, NS3, and hA3G protein (or APOBEC proteins with HA tag) was detected with western blot. A similar procedure was used for the experiment using GS4.3 cells, except the infection step. Huh7.5 cells were planted into the 6-well plate with 3 × 105 cells per well in the complete growth medium and infected with HCV viral stock (45 IU per cell).

Eradication rates with standard triple therapy, which originally achieved 90% eradication, are now being observed to be consistently lower than 70–80% [4,27,28]. Studies published over the last 2 years have

compared some of the therapies which had hitherto been more commonly used as second and subsequent line therapies with standard triple therapy. Cytoskeletal Signaling inhibitor One such trial compared a 10 -day bismuth-based regime containing metronidazole, tetracycline and omeprazole (OBMT) against a 7 -day course of triple therapy with omeprazole, amoxicilin, and clarithromycin (OAC) and found a superior eradication rate among the OBMT group. Eradication rates were 93.3% with OBMT and 69.6% with OAC in the per-protocol population (p < .001) and 79.8% and 55.4%, respectively in the ITT population. As encouraging

as these results are, it still falls short of the 80% eradication rate based on ITT which is desirable under the Maastricht consensus [28]. A criticism of this trial has been that it ought to have compared the OBMT regimen with a 10 -day OAC regime. It has also been postulated that longer treatment durations for bismuth-based therapy may be more efficacious. A study that looked at a 14 -day OBMT selleck chemicals llc regime in a mixture of first line and salvage treatments showed an eradication rate of 95% by find more ITT analysis [29]. Therefore, the optimum duration of OBMT treatment is not yet clear. Levofloxacin may also have a role to play as a first-line treatment strategy. In light of the increase in clarithromycin resistance, one trial looked at substituting levofloxacin for clarithromycin in both standard triple and sequential regimes, all on a 10 -day basis. Of the four treatment arms, levofloxacin consistently outperformed clarithromycin in sequential and standard therapies with the best results coming from the sequential arm

that contained levofloxacin, which had an ITT eradication rate of 82.5% [22]. The optimal duration of treatment for all forms of H. pylori eradication treatment is tending toward longer courses, and this has been discussed in the Maastricht and ACG guidelines [5,25]. The topic is currently the subject of a Cochrane review and is at the protocol stage. Bismuth and levofloxacin-based therapies are very frequently used as second-line therapies also, a setting in which their efficacy is long acknowledged. Bismuth-based therapy has an efficacy of 76% in second-line therapy on the basis of a pooled analysis [30]. It is also safe with no serious side effects reported in a cohort of 4763 patients receiving it for H. pylori eradication [31]. Some case reports have suggested a risk of black tongue [32]. Other bismuth-based treatment regimes have been proposed for second-line therapy.