Interestingly, mitochondria, nuclei, and endoplasmic reticulum remained morphologically unchanged. Cholesterol and neutral lipids (TG and cholesterol esters) were quantified by way of gas/liquid chromatography (Fig. 3). Whereas tetracycline caused no significant changes in TG content after 24 hours,

a six-fold increase was induced by a 50 μM concentration after 14 days. By contrast, selleck cholesterol and cholesterol esters content remained unchanged. A dose-dependent increase in TG content was also observed, and cholesterol esters were slightly augmented in HepaRG cells treated by amiodarone for 14 days. In addition, phospholipids (phosphatidylethanolamine, phosphatidylcholine, sphingomyelin, phosphatidylserine, and phosphatidylinositol) were measured by way of HPLC in HepaRG cells treated with 20 μM amiodarone for 24 hours or 14 days (Fig. 4). Whereas no significant change was observed in phospholipid content after acute exposure, phosphatidylethanolamine and phosphatidylcholine levels were strongly enhanced, and sphingomyelin, phosphatidylserine, and phosphatidylinositol levels were slightly augmented after 14 days. Impairment of mitochondrial fatty acid oxidation (FAO) is considered one of the major mechanisms of liver steatosis.21 FAO was evaluated by measuring [14C]-labeled acid-soluble

β-oxidation products in HepaRG cells after 24-hour and 14-day treatments using either 20 μM tetracycline or 50 μM amiodarone (Fig. 5). A 20% diminution of FAO was observed after both acute and chronic CP868596 amiodarone treatments, and only after chronic tetracycline exposure. To characterize gene expression changes associated with induction of phospholipidosis and steatosis, the transcriptome of HepaRG cells was analyzed after 24-hour and 14-day treatments with 20 μM amiodarone using pangenomic

oligonucleotide microarrays. Significantly modulated genes were extracted with a fold change >1.5 or <−1.5 and P ≤ 0.01 as filters. Their total numbers reached 547 and 594 with up-regulated genes representing 48% and 44%, after 24-hour and 14-day exposure, respectively (Supporting Tables 1 and 2); 176 genes were in common at the two time points. Functional analysis revealed that expression of many genes involved in the regulation of lipid metabolism (including ACOT12, ADFP, ALDH3A1, APOA2, FASN, MOGAT1, SREBP1, learn more and THRSP) or related to phospholipidosis (such as LSS, LPIN1, ASML3A, and GDPD3) was significantly altered. Various genes regulating growth/proliferation, cell death, assembly/organization, and inflammation were also substantially deregulated. To validate and complete this microarray analysis, changes in the expression of 29 genes, which are key players in lipid metabolism and/or liver-specific functions, were further examined by way of RT-qPCR in HepaRG cells exposed to several concentrations of amiodarone (5-20 μM), tetracycline (10-100 μM), and oleic acid (100-500 μM) for 24 hours or 14 days. The data are displayed in Table 2.

We divide the therapeutic approach to PDPH into 4 stages: conservative management, aggressive medical management, conventional invasive treatments, and the very rarely employed less conventional invasive treatments and provide management algorithm to facilitate treatment. “
“Objectives.— Migraine is a risk factor for stroke in young women. Biomarker studies implicate endothelial activation as a possible mechanism. Emerging Roxadustat concentration relationships of childhood adversity with migraine, and with inflammation, a component

of endothelial activation, suggest that it may play a role in the migraine–stroke association. Our objective is to evaluate the relationship between adverse childhood experiences (ACEs), migraine, and vascular biomarker levels in premenopausal women. Methods.— Vascular and metabolic biomarkers from women 18-50 years, including 125 with migraine (interictal) and 50 without migraine, were evaluated. An ACE questionnaire was later collected by mail (response rate 80.6%, 100 migraineurs, 41 controls). Results.— Migraineurs

and controls were demographically similar. Migraineurs reported adversity more commonly than controls (71% vs 46%, odds ratio [OR] = 1.53, 95% confidence interval 1.07-2.17). Average ACE scores were elevated in migraineurs as compared with controls (2.4 vs 0.76, P < .001). STA-9090 in vitro ACE scores correlated with headache frequency (0.37, P = .001) and younger age of headache onset (−0.22, P = .04). It also correlated with body mass index (r = 0.43, P = .0001), von find more Willebrand factor activity (r = 0.21, P = .009), tissue plasminogen activator antigen (r = 0.28, P = .004), prothrombin

activation fragment (r = 0.36, P = .001), high-sensitivity C-reactive protein (r = 0.98, P = .0001), transforming growth factor-beta1 (r = 0.28, P = .003), tissue necrosis factor-alpha (r = 0.20, P = .03), interleukin-6 (r = 0.22, P = .03), adiponectin (r = −0.29, P = .003), and nitrate/nitrite concentration (r = −314, P = .001). Logistic regression analyses (adjusted for vascular risk factors and migraine) demonstrated an association of childhood adversity with inflammatory factors (high-sensitivity C-reactive protein, interleukin-6, and tissue necrosis factor-alpha). Conclusions.— In young women, adverse childhood events are associated with migraine, particularly chronic and transformed migraine, and with vascular biomarkers, especially inflammatory biomarkers. These findings implicate early life stress as a link between migraine and endothelial activation. “
“(Headache 2010;50:1005-1016) Objective.— To investigate the functional abnormalities of the motor cortices in children with migraine using magnetoencephalography (MEG) and a finger-tapping task. Background.— Cortical hyperexcitability has been reported in adults with migraine using MEG. Many children with migraine report difficulty with motor functioning.

The authors do not elaborate on other possible mechanisms, but given the rapidity of the effect whereby the LSECs induce a maximal proliferative response in the hepatocytes within 2 days of hepatectomy, one possibility

is that Daporinad cell line it acts through a stress response that is induced through the large reduction in liver function which must accompany a 70% reduction in liver size. Indeed, there are a number of metabolic sensors that activate an angiogenic response,4 with many acting through VEGF. Previous studies have shown a significant up-regulation of VEGF5 within 24 hours after hepatectomy, and because the LSECs express VEGFR2, one could postulate this results in activation of the LSECs, leading to the release of signals that result in hepatocyte proliferation. In support of the idea that LSECs are activated through a stress response pathway is the spectrum of genes identified as being altered in the regenerative liver, a large number of which are involved in a stress response leading to a DNA damage response. Genes such as RAD51 and RAD54 (RecA homologs, E. coli) and their associated proteins were highly up-regulated in the array. These are members of genes associated

with machinery for the detection and repair of DNA double-stranded breaks, part of the DNA damage response. Also up-regulated were a number of S100 binding proteins, proteins known to regulate inflammation and to confer protection from oxidative damage. Hydroxychloroquine In addition, a large number of genes involved in cell cycle control were highly increased. Finally, this study demonstrates the requirement for LSEC–hepatocyte interactions in the response to injury. In general, the endothelium is heavily dependent on juxtacrine signals (i.e., ones delivered by direct cell contact) in contrast to the endocrine and hematopoietic systems. These signals can be delivered through cells such as smooth muscle cells, pericytes, glial cells in the brain, or astrocytes in the eye. In the liver, previous studies have shown that the maintenance of

the specialized phenotype of both the LSEC and the hepatocyte requires cell-to-cell contact. Isolated LSECs show a loss of their characteristic find more features within 24 hours, such as rounded cell shape and fenestrations, indicating that normal LSEC differentiation in vivo depends on the hepatic microenvironment, the nature of which has not been adequately addressed. Three-dimensional coculture also improves hepatocyte function.6 The study further shows that cellular contact between the activated LSECs and hepatocytes is also essential for the induction of proliferation of the hepatocytes. Wnt2 (Wnts are secreted factors that act through the Frizzled receptors and are involved in cell fate determination and progenitor cell proliferation) and hepatocyte growth factor were identified as crucial factors in driving the proliferative response of hepatocytes.

Distinctions in habitat occupation can be recognized at various scales, from landscape regions through vegetation types to the composition of local resource patches (Senft et al., 1987). Differences in food resources can be identified at plant species level, or in terms of the local accessibility and nutritional value of plant parts or growth stages. Relevant features of resource heterogeneity at landscape scale include woody vegetation structure (Ferrar & Walker, 1974; Greenacre & Vrba, 1984) and soil fertility (East, 1984). Choice

of feeding sites and plant species and parts consumed by herbivores Kinase Inhibitor Library datasheet is influenced by nutrient and fibre contents (Ben-Shahar & Coe, 1992; Bailey et al., 1996),

dependent on grass height and relative greenness (Wilmshurst et al., 1999). Hence, coexistence among large herbivores may be enabled by distinctions in resource use at one or more of these scales, underlain by differences in body size and morphological adaptations. Sable antelope GPCR Compound Library high throughput Hippotragus niger are medium-sized ruminants (adult female body mass 220 kg) with relatively narrow muzzles (incisor arcade breadth 57 mm; Gordon & Illius, 1988) that enable them to graze tall grass (Skinner & Chimimba, 2005). Their highest recorded density is about three animals per km2 (Grobler, 1974), but their local abundance within the Kruger National Park (KNP) where our study area was located did not exceed 0.5 animals per km2 (Chirima et al., unpubl. data). African buffalo (Syncerus caffer) are

large ruminants (adult female body mass 520 kg) with broad muzzles (incisor arcade breadth 93 mm), and are bulk grazers on tall grass. They attain a regional this website density of 1.5 animals per km2 within KNP, and local densities of 3–5 animals per km2. Plains zebra (Equus quagga) are medium-large non-ruminants (adult female body mass 310 kg) that tolerate quite tall and hence fibrous grass due to their hindgut digestion. They exhibit local and regional densities in KNP closely similar to those of buffalo. Differences in social groupings may also influence resource exploitation patterns. Buffalo form large herds generally numbering several hundred animals and zebra cohesive groups of 5–10 animals, while sable herds typically number 15–30 animals (Grobler, 1974; Sinclair, 1977; Skinner & Chimimba, 2005).

Distinctions in habitat occupation can be recognized at various scales, from landscape regions through vegetation types to the composition of local resource patches (Senft et al., 1987). Differences in food resources can be identified at plant species level, or in terms of the local accessibility and nutritional value of plant parts or growth stages. Relevant features of resource heterogeneity at landscape scale include woody vegetation structure (Ferrar & Walker, 1974; Greenacre & Vrba, 1984) and soil fertility (East, 1984). Choice

of feeding sites and plant species and parts consumed by herbivores www.selleckchem.com/products/Erlotinib-Hydrochloride.html is influenced by nutrient and fibre contents (Ben-Shahar & Coe, 1992; Bailey et al., 1996),

dependent on grass height and relative greenness (Wilmshurst et al., 1999). Hence, coexistence among large herbivores may be enabled by distinctions in resource use at one or more of these scales, underlain by differences in body size and morphological adaptations. Sable antelope MK0683 supplier Hippotragus niger are medium-sized ruminants (adult female body mass 220 kg) with relatively narrow muzzles (incisor arcade breadth 57 mm; Gordon & Illius, 1988) that enable them to graze tall grass (Skinner & Chimimba, 2005). Their highest recorded density is about three animals per km2 (Grobler, 1974), but their local abundance within the Kruger National Park (KNP) where our study area was located did not exceed 0.5 animals per km2 (Chirima et al., unpubl. data). African buffalo (Syncerus caffer) are

large ruminants (adult female body mass 520 kg) with broad muzzles (incisor arcade breadth 93 mm), and are bulk grazers on tall grass. They attain a regional selleck products density of 1.5 animals per km2 within KNP, and local densities of 3–5 animals per km2. Plains zebra (Equus quagga) are medium-large non-ruminants (adult female body mass 310 kg) that tolerate quite tall and hence fibrous grass due to their hindgut digestion. They exhibit local and regional densities in KNP closely similar to those of buffalo. Differences in social groupings may also influence resource exploitation patterns. Buffalo form large herds generally numbering several hundred animals and zebra cohesive groups of 5–10 animals, while sable herds typically number 15–30 animals (Grobler, 1974; Sinclair, 1977; Skinner & Chimimba, 2005).

In response to TAT-ARC pretreatment, survival was significantly improved in both PLX4032 chemical structure models compared to PBS or TAT-βgal-pretreated hepatocytes. TAT-ARC-mediated protection of hepatocytes was in both models comparable to that of JNK-inhibitor SP600125 pretreated hepatocytes (Fig. 5A). TNF/AcD or TNF/GalN stimulation of hepatocytes resulted in JNK activation that was inhibited by TAT-ARC or JNK-inhibitor pretreatment (Fig. 5B). In both models, ConA- and GalN/LPS-induced hepatitis, TNF-α levels have been shown

to be critical for hepatocyte killing and high mortality of the animals. Thus, we examined the effect of TAT-ARC on serum TNF-α levels in murine models of hepatitis caused by ConA and GalN/LPS. Importantly, in both models TAT-ARC significantly reduced serum TNF-α levels (Fig. 5C). Together, these data suggest that TAT-ARC prevents TNF-mediated hepatitis by inhibiting TNF-α expression, e.g., in nonparenchymal cells, but also directly protects hepatocytes from apoptosis. The crucial role of JNK signaling in TNF-dependent ALF

was demonstrated by Maeda et al.,21 who showed that mice lacking either JNK1 or JNK2 are highly resistant to ConA-induced ALF. Thus, we investigated JNK activation by immunoblot analysis using liver lysates from both ConA- and GalN/LPS-treated mice. As shown in Fig. 6A, both p46- and p54-JNK phosphorylation, which are essential steps for JNK activation, were significantly Z-VAD-FMK manufacturer induced after ConA or GalN/LPS stimulation. Interestingly, both p46- and p54-JNK phosphorylation were strongly reduced in TAT-ARC-treated mice following ConA stimulation and were completely abrogated after GalN/LPS administration (Fig. 6A). Because activated JNK translocates from cytosol to mitochondria to trigger cell death JNK translocation was assessed by subcellular fractionation and immunoblotting of liver lysates.22 TAT-ARC application completely blocked JNK activation

and subsequent mitochondrial translocation selleck chemicals llc following ConA or GalN/LPS, respectively (Fig. 6B). Because the death-promoting function of JNK-signaling in the liver is antagonized by p38 signaling, p38α phosphorylation was analyzed23 (Fig. 6A). Although no relevant activation of p38-signaling was detected 4 hours after GalN/LPS stimulation when JNK was already activated, TAT-ARC-mediated hepatoprotection following ConA stimulation was associated with a substantial concomitant activation of p38α signaling. It remains unclear whether p38α activation following ConA and TAT-ARC treatment plays a causal role for the observed protective effect seen or is rather a secondary phenomenon. JNK specifically regulates the proapoptotic activity of BH3-only proteins Bax and Bim, which cooperate with Bid in hepatocyte killing.

HEV cases were matched by year and transplant type to negative controls in a 1:3 ratio, and assessed by Chi square and multivariable conditional logistic regression. Results: Of 311 subjects (271 kidney,

33 lung, 5 heart, 2 liver) in our cohort, 16 (13 kidney, 2 lung, 1 liver) demonstrated evidence of post-transplant HEV infection (4 by HEV PCR, 2 by anti-HEV IgM,10 by anti-HEV IgG seroconversion) and were matched to 48 controls. Univariate analysis revealed significant associations between post transplant HEV infections, cyclosporine use (p=0.015), and leukopenia (p=0.007). In the multivariable model, leukopenia (OR 4.15), thrombocytopenia (OR 2.24) and tacrolimus use (OR 1.09) were associated with increased risk of HEV infection post SOT, though only leukopenia was statistically significant (p=.04). No subjects developed chronic HEV infection. Conclusions: Leukopenia was associated selleck kinase inhibitor with an increased risk of post-transplant HEV infection in our cohort. Associations with other variables suggest a relationship between

immunosuppression and risk of infection, but were not statistically significant. In contrast to previous studies, we did not identify any chronic HEV infections. Our findings suggest that while important, immunosuppression and exposure alone may be insufficient for the establishment of chronic HEV infection among SOT recipients. Disclosures: Kathleen B. Schwarz – Consulting: Novartis, Novartis; Grant/Research Support: Bristol-Myers Squibb, Gilead, Roche/Genentech, Bristol-Myers Squibb, Vertex, Roche The following people have nothing Belnacasan cell line selleck chemical to disclose: Paul K. Sue, Nora Pisanic, Christopher D. Heaney, Kenrad Nelson, Alexandra Valsamakis, Michael Forman, Annette M. Jackson, John R. Ticehurst, Robert A. Montgomery, Wikrom Karnsakul Histological recurrence of hepatitis C (HCV) post-liver transplantation (LT) is still an important event even in the era of more effective HCV treatments. The Hepatitis Aggressiveness Score (HAS) is a histologic classification system that has been

recently developed to assess the recurrence of HCV. Objective: the main outcome of the study was to evaluate graft survival time based on HAS and to assess pathologist inter-observer agreement. Methods: we reviewed the clinical records of HCV liver transplant recipients in our facility from June 1999 to June 2012. We included those patients who had >30 day survival. Clinical and histologic characteristics were obtained. Biopsies were independently evaluated by 3 pathologists. All biopsies were assessed for the presence of the following features, which comprise the basis of the HAS: 1) prominent ductular reaction 2) prominent hepatocyte ballooning 3) cholestasis (including at least focal canalicular cholestasis of any degree) and 4) periportal sinusoidal/pericellular fibrosis.

A promoter assay and western blotting confirmed that peretinoin promoted the autophagy GSK126 manufacturer related-gene Atg16L1, and electron microscopy revealed increased autophagosome formation due to peretinoin. Atg16L1-transfected HepG2 cells showed suppressed expression of pStat3 and pNFkB. Recombinant IL-6 and palmitic acid induced expression of pSTAT3 and pNFkB in primary hepatocytes in vitro, whereas peretinoin dose-de-pendently suppressed the expression of these genes. Conclusion Peretinoin suppresses the

development of liver steatosis, inflammation, and tumorigenesis by activating Atg16L1-depen-dent autophagy. These results support the clinical efficacy of peretinoin for preventing HCC. Disclosures: Hikari Okada – Employment: Kanazawa University Shuichi Kaneko – Grant/Research Support: MDS, Co., Inc, Chugai Pharma., Co., Inc, Toray Co., Inc, Daiichi Sankyo., Co., Inc, Dainippon Sumitomo, Co., Inc, Ajinomoto Co., Inc, MDS, Co., Inc, Chugai Pharma., Co., Inc, Toray Co., Inc, Daiichi Sankyo., Co., Inc, Dainippon Sumitomo, Co., Inc, Ajinomoto Co., Inc, Bayer Japan The following people have nothing to disclose: Masao Honda, Pexidartinib research buy Kai Takegoshi, Naoto Matsuzawa, Taro Yamashita, Yoshio Sakai, Toshinari Takamura, Takuji Tanaka Background and Aims: Cholangiocarcinoma (CCA) is a lethal

neoplasm originating from the biliary epithelium. Risk factors for CCA include liver inflammation and fibrosis. IL-33 has been shown to promote liver inflammation and fibrosis. The AKT cell survival and Hippo cell growth controlling pathways are involved in CCA carcinogenesis and progression. Yes-associated protein (YAP) is a major transcriptional factor of the Hippo pathway. Our aim was to generate a mouse model of CCA incorporating these oncogenic pathways mimicking the human disease. Methods: Ectopic oncogene expression in the biliary tract was accomplished by the Sleeping Beauty transposon transfection system with transduction of constitutively

active AKT and/or YAP. Intrabiliary injection of the transposon-transposase complex was coupled with lobar bile duct ligation in CL57/BL6 wild type or IL-6 −/− mice. After injection, mice were treated with either vehicle or IL-33 i.p. for three consecutive selleck chemicals llc days. Mice were sacrificed on day 70 and examined for the presence of tumors and tumor burden. Results: Intrabiliary instillation of a sleeping beauty transposon-transposase complex expressing GFP revealed significant transduction of the cholangiocytes, but not hepatocytes, in the bile duct ligated lobe 7 days later; GFP transduction did not require IL-33 administration. Tumor development following intrabiliary instillation of AKT plus YAP, however, was augmented in animals treated with IL-33. Tumors developed in 60% of the animals receiving both oncogenes plus IL-33, but in only 20% of the mice transduced with the oncogenes alone (p<0.05).

The same quantity MK-8669 chemical structure of total RNA was reverse-transcribed to complementary DNA (cDNA)

using M-MLV Transcriptase (Invitrogen) in the presence of oligo-dT primers (Shenggong, China). Quantitative PCR was performed using SYBR Green I (Takara) for 45 cycles at 15 seconds at 95° and 60 seconds at 60° with Rotor-Gene 6000 (Corbett Research) according to the manufacturer’s instructions. Quantitative PCR primers were included in Supporting Information materials. Results were analyzed by ΔΔCt method as described before.28 Values were expressed as fold change in comparison with control. Donor splenocytes were isolated from biweekly CCl4-treated C57BL/6 and HBV-tg mice. Four million splenocytes were adoptive transferred weekly (i.p.) to Rag1−/− recipient mice from PD98059 cell line the same genetic background and age for 4 weeks. The recipient Rag1−/− mice were

sacrificed and liver tissues were collected 72 hours after the fourth transfer. HSCs were isolated from liver of HBV-tg mice by collagenase and pronase perfusion and 8.2% Nycodenz (Sigma) gradient centrifugation.29 For isolation of hepatic NKT cells, first, HBV-tg mice were injected CCl4 (i.p. 0.5 μL of body weight) 12 hours and isolation of liver mononuclear cells (MNCs). Liver MNCs were stained for PE-NK1.1 and APC-CD3 (BD PharMingen, San Diego, CA) and sorted by flow cytometry (Becton Dickinson) for CD3+NK1.1+ NKT cells. For the coculture experiment, HSCs were previously cultured for 24 hours before constitution of NKT cells, and then cocultured (1:10) with CCl4-pretreated NKT cells for another 24 hours with or without functional purified neutralizing cytokine antibodies of IL-4, IL-13, or IFN-γ at a concentration of 5 μg/mL (eBioscience).

After coculture, NKT cells were removed by washing, HSCs were visualized with phase-contrast microscopy, and collected by mild trypsinization check details for analyzing the transcription of α-SMA. HSCs RNA extraction were using RNAprep pure Micro Kit (Tiangen Biotech, Beijing, China). Analysis of liver transaminase activity, liver histology, and immunohistochemistry for α-SMA, liver mononuclear cell (MNC) isolation and flow cytometric analysis, cell depletion, NKT cell preparation, and adoptive transfer to Rag1−/− mice in vivo CD1d block methods are included in the Supporting Information Materials online. Student t test was chosen to compare values between two groups. Analysis of variance (ANOVA) was used to compare values from multiple groups. Data are expressed as means ± standard deviation (SD). P < 0.05 was considered statistically significant. It was found that HBV-tg mice had an elevated level of serum ALT at the age of 2, 3, 4, and 6 months than that of normal C57BL/6 mice, showing that ALT levels were much higher in HBV-tg mice compared with C57BL/6 mice (all below 40 IU/L) (Fig.

5, 6 A key requirement for maintaining long-term HBV DNA suppression is the avoidance of resistance to the antivirals. The current cohort analysis followed entecavir-treated patients from ETV-022 who continued to receive entecavir in ETV-901 through up to 5 years of therapy. Most patients who did not enter ETV-901 were responders in ETV-022, the majority of whom achieved HBV DNA <300 copies/mL through 2 years of entecavir therapy.19 Despite high proportions of the previous nonresponders and only one previous responder in the entecavir long-term cohort, high rates of HBV DNA to <300 copies/mL (94%) and normal

ALT levels (80%) were achieved or maintained at 5 years. Patients in ETV-901 received entecavir 1.0 mg daily and in some cases had a period of treatment with both entecavir and lamivudine. Most patients in the entecavir long-term ICG-001 purchase cohort with HBV DNA <300 copies/mL had already achieved this endpoint after 2 years of treatment with entecavir 0.5 mg daily. This result suggests that the dosage increase

from 0.5 mg (in ETV-022) to Romidepsin manufacturer 1.0 mg (in ETV-901) had minimal contribution to the virologic suppression noted in the long-term cohort. Further evidence that entecavir at a dose of 0.5 mg daily achieves and maintains HBV DNA suppression during long-term therapy is suggested by a recent study of entecavir in Japanese patients with CHB. In that study, 87% of patients achieved HBV DNA <400 copies/mL after 3 years of continuous treatment with entecavir 0.5 mg daily.26 Treatment this website with entecavir beyond 2 years resulted in incremental benefits for serologic response. Most patients who had previously undergone HBeAg seroconversion after 2 years of entecavir therapy in study ETV-022 were categorized as responders and did not enroll in study ETV-901. Therefore, HBeAg

seroconversion occurring in 23% (33/141) of the entecavir long-term cohort during study ETV-901 represents an incremental improvement in serologic response in a difficult-to-treat population, in addition to the 31% of patients with HBeAg seroconversion through 2 years in study ETV-022.19 Continued treatment with entecavir beyond 2 years also resulted in incremental benefit of HBsAg loss: in addition to the 18 (5%) patients who lost HBsAg during ETV-022, two more patients (1.4%, 2/145) in the entecavir long-term cohort lost HBsAg during study ETV-901. It should be noted that HBV serology assays during study ETV-901 were performed at local laboratories using variable methodologies, in contrast to the uniform assay performed in a central laboratory during study ETV-022. Patient drop-out and missing data are common during long-term studies. In this study, 47 patients had discontinued treatment prior to Year 5, and five patients who were still on-treatment at Year 5 had missing HBV DNA measurements.