05) (Supporting Fig. 6). In contrast, IL-10 was significantly lower in mice that received Con PD0325901 A only (P < 0.05), whereas there was no significant difference in the serum level of TNFα between experimental groups. We also examined whether TD139 would affect the infiltration of MNCs into the livers of

mice after Con A injection (Fig. 7). Although there was no significant difference in the total number of liver-infiltrating MNCs, CD3+ T cells, NK1.1+ NK and CD3+NK1.1+ NKT cells, CD11c+ DCs and CD11c+CD80+CD86+ activated DCs, F4/80+ macrophages (data not shown), and Gal-3 INH profoundly reduced CD4+- and CD8+ T-cell infiltration into the liver parenchyma (Fig. 7). The total number of IFNγ- and IL-17- and -4-producing CD4+ T cells and IFNγ-producing CD8+ T lymphocytes was significantly lower in TD139-treated mice (Fig. 7). However, HM781-36B cost the total number of CD4+ IL-10-producing cells was significantly higher in TD139-treated mice (P < 0.05). There was no difference in the total number of TNFα- producing CD4+ T cells, IFNγ- and IL-4-producing CD3+NK1.1

NKT cells, IFNγ-producing NK1.1+ NK cells, IL-12- and -10-producing CD11c+ DCs, and F4/80+ macrophages between TD139-treated mice and mice that received Con A only (data not shown). In line with results obtained from Gal3−/− mice (Fig. 5A), we found a significantly higher percentage and total number of F4/80+ CD206+ alternatively activated (i.e., M2-polarized) macrophages in livers of TD139-treated mice, compared to mice treated with Con A only (Fig. 8A). However, there was no difference in the total number of IL-12- and -10-producing macrophages (data not shown). We also found that TD139 prevented apoptosis of liver-infiltrating MNCs (Fig. 8B) 8 hours after Con A injection. Significantly lower percentages of Annexin V+ PI+ liver-infiltrating MNCs (P < 0.01) were observed in TD139-treated mice, compared to mice treated with Con A only. Here, we provided

the first evidence that Gal-3 plays an important role in the pathogenesis of Con A–induced hepatitis, a model of acute, fulminate hepatitis in humans.1 Targeted disruption of Gal-3 gene attenuated liver injury by reducing the number of effector cells, including T lymphocytes (both CD4+ and CD8+), B lymphocytes, DCs, and NK and NKT cells, and increasing the number of IL-10-producing CD4+ T cells selleck screening library and alternatively activated (i.e., M2-polarized) macrophages that was accompanied with lower serum levels of TNFα, IFNγ, and IL-17, but also IL-4. In addition, pretreatment of WT mice with TD139 attenuated Con A–induced liver injury (Fig. 6). IP injection of TD139 in WT mice 2 hours before and immediately after Con A injection suppressed the infiltration of IFNγ- and IL-17- and -4-producing CD4+ and IFNγ-producing CD8+ lymphocytes (Fig. 7), increased the total number of IL-10-producing CD4+ T cells (Fig. 7), attenuated serum levels of IFNγ and IL-17 and -4, elevated the serum level of IL-10 (Supporting Fig.

Further, greater CPM waning in the CPM-sequential series was correlated with less reported extent of pain reduction by episodic medication (r = 0.493, P = .028). Migraineurs have subtle deficits in endogenous

pain modulation which requires a more challenging test protocol than the commonly used single CPM. Waning of CPM response seems to reveal this pronociceptive state. The clinical relevance of the CPM waning effect is highlighted by its association with clinical parameters of migraine. “
“Prion protein, a sialoglycoprotein with neuroprotective properties on oxidative stress damage, has been related with the mechanisms leading to migraine. In the present case-control study, we investigated the correlation between the common methionine/valine polymorphism at codon 129 within the prion protein gene (PRNP) and migraine. Genotyping of PRNP V129M variant was performed in 384 migraine patients and 185 age-, sex-, and race-ethnicity-matched GSK126 chemical structure healthy controls. The frequencies of the PRNP V129M genotype did not differ significantly between

migraineurs and controls. The frequencies of 129VV genotype were significantly higher in patients with earlier age at migraine onset. No correlation was found between PRNP 129 genotype and demographics, and PD-0332991 mw other clinical migraine features. Our data suggest that the PRNP 129VV polymorphism is not a direct migraine risk factor but is significantly associated with an earlier onset of the disease. “
“Objective.— To investigate the alteration of hippocampal long-term plasticity and basal synaptic transmission induced selleck chemical by repetitive cortical spreading depressions (CSDs). Background.— There is a relationship between migraine aura and amnesia attack. CSD, a state underlying migraine attacks, may be responsible for hippocampus-related symptoms. However, the precise role of CSD on hippocampal activity has not been investigated. Methods.— Male Wistar rats were divided into CSD and control groups. Repetitive CSDs were induced in vivo by topical application of solid KCl.

Forty-five minutes later, the ipsilateral hippocampus was removed, and hippocampal slices were prepared for a series of electrophysiological studies. Results.— Repetitive CSDs led to a decrease in the magnitude of long-term potentiation in the hippocampus. CSD also reduced hippocampal synaptic efficacy, as shown by a reduction in post-synaptic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor responses. In contrast, the post-synaptic N-methyl-d-aspartate receptor responses remained unchanged. In addition, there were no changes in paired-pulse profiles between the groups, indicating that CSD did not induce any presynaptic alterations. Conclusion.— These findings suggest that a reduction of post-synaptic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor responses is the mechanism responsible for impaired hippocampal long-term potentiation induced by CSD.

Included were 133 adult and paediatric patients with a high suspicion of VWD. Fifty-three were diagnosed with VWD: 47 (88.7%) with type 1 VWD, four (7.5%) with type 2a VWD and two (3.8%) with type 3 VWD. Mean age for

female patients was 19.5 years (range 3–44 years) and 18.5 years (range 4–63 years) for male patients. Mean age at start of bleeding symptoms was 8.8 years (range 1–61). The most frequent clinical symptoms were epistaxis (84.9%), ecchymosis (79.2%), haematomas (71.7%), gum bleeds (62.3%) and petechia (50.9%). Severe transoperative or postoperative bleeding was found in 17 patients (32.1%). Twenty-six women at childbearing age had a history of abnormal gynaecological bleeding. Our results clearly demonstrate the presence of VWD in Mexican and underscore the importance of a more detailed description of VWD. DAPT supplier Efforts to increase the awareness and diagnosis of VWD could help in better identification of patients with bleeding disorders and lead to early, appropriate management with safe and efficacious therapies such as desmopressin and

plasma concentrates. “
“Joint bleeding is the hallmark of severe haemophilia and the major cause of disability in patients with this coagulopathy. Repeated bleeding into the same AZD1208 datasheet joint can lead to chronic synovitis and progressive arthropathy. Radiosynovectomy is one option for the treatment of chronic haemophilic synovitis, but concerns about the risks of exposure

to ionizing radiation have divided clinicians as to the safety and appropriate use of the procedure. This article presents two differing viewpoints, one from a pair of orthopaedic surgeons who collectively have performed more than 300 radiosynovectomies in patients with haemophilia. They maintain that radiosynovectomy is a simple, effective, safe and low-cost technique children and adults with chronic haemophilic synovitis. The other perspective is from an experienced haemophilia treater who directs a major US haemophilia treatment centre. She believes that unresolved questions about the safety of radiation exposure in children argue against the use of radiosynovectomy in paediatric patients with haemophilia. “
“Summary.  Progress in the evidence-based care of haemophilia A and B worldwide has this website been historically challenged by the dearth of evaluable outcome data, including but not limited to the safety and effectiveness of therapeutic interventions. These challenges are partially rooted in the inherent difficulty of conducting prospective clinical trials and observational studies with statistically meaningful endpoints in a rare disease such as haemophilia. Despite the logistical barriers, the need for outcome data has never been more critical than in this time of expansive therapeutic advance tempered by the shrinking economic capacity to fund the rapidly increasing cost of treatment.

Included were 133 adult and paediatric patients with a high suspicion of VWD. Fifty-three were diagnosed with VWD: 47 (88.7%) with type 1 VWD, four (7.5%) with type 2a VWD and two (3.8%) with type 3 VWD. Mean age for

female patients was 19.5 years (range 3–44 years) and 18.5 years (range 4–63 years) for male patients. Mean age at start of bleeding symptoms was 8.8 years (range 1–61). The most frequent clinical symptoms were epistaxis (84.9%), ecchymosis (79.2%), haematomas (71.7%), gum bleeds (62.3%) and petechia (50.9%). Severe transoperative or postoperative bleeding was found in 17 patients (32.1%). Twenty-six women at childbearing age had a history of abnormal gynaecological bleeding. Our results clearly demonstrate the presence of VWD in Mexican and underscore the importance of a more detailed description of VWD. XL765 nmr Efforts to increase the awareness and diagnosis of VWD could help in better identification of patients with bleeding disorders and lead to early, appropriate management with safe and efficacious therapies such as desmopressin and

plasma concentrates. “
“Joint bleeding is the hallmark of severe haemophilia and the major cause of disability in patients with this coagulopathy. Repeated bleeding into the same Roxadustat clinical trial joint can lead to chronic synovitis and progressive arthropathy. Radiosynovectomy is one option for the treatment of chronic haemophilic synovitis, but concerns about the risks of exposure

to ionizing radiation have divided clinicians as to the safety and appropriate use of the procedure. This article presents two differing viewpoints, one from a pair of orthopaedic surgeons who collectively have performed more than 300 radiosynovectomies in patients with haemophilia. They maintain that radiosynovectomy is a simple, effective, safe and low-cost technique children and adults with chronic haemophilic synovitis. The other perspective is from an experienced haemophilia treater who directs a major US haemophilia treatment centre. She believes that unresolved questions about the safety of radiation exposure in children argue against the use of radiosynovectomy in paediatric patients with haemophilia. “
“Summary.  Progress in the evidence-based care of haemophilia A and B worldwide has selleck kinase inhibitor been historically challenged by the dearth of evaluable outcome data, including but not limited to the safety and effectiveness of therapeutic interventions. These challenges are partially rooted in the inherent difficulty of conducting prospective clinical trials and observational studies with statistically meaningful endpoints in a rare disease such as haemophilia. Despite the logistical barriers, the need for outcome data has never been more critical than in this time of expansive therapeutic advance tempered by the shrinking economic capacity to fund the rapidly increasing cost of treatment.

This is also true for the recently licensed protease

inhibitors, which I-BET-762 solubility dmso markedly improved SVR rates in patients with CHC, GT-1.9-14 As far as we know, this is the first study to assess a potential selection bias in patients with CHC treated within randomized, controlled studies. At least, the outcome of cohort studies in hepatitis C may be influenced by a selection bias.21 Not surprisingly, some baseline characteristics, adherence to treatment, and the IL18B phenotype had the biggest effect on treatment outcome. Patients receiving SOC had more advanced liver disease, a more frequent history of psychiatric disorders, and a substantial proportion was on drug-substitution therapy. Differences in baseline characteristics22-25 reflect inclusion and exclusion criteria and may create “perfect to treat” patients. Whether the slight differences documented in this study are clinically relevant remains unknown, but history of psychiatric disorders, need of drug-substitution therapy, and a higher grade of liver fibrosis may influence treatment outcome, as well. Because of major differences in access to

selleck medical care, data from a European study center cannot be directly applied to the U.S. population of HCV patients. It may also be difficult to draw conclusions from the largely homogenous population of HCV patients seen in Vienna to the largely heterogeneous population of HCV patients seen in the United States. In particular, in this study, except for 7 patients (2.3%; 5 Asians and 2 black Africans), all patients were Caucasians, whereas in the United States, approximately 19% and 40% of treatment eligible or noneligible

patients, respectively, are black.26 Analysis of data from the National Health and Nutrition Examination Survey, conducted in 2005-2008, revealed that HCV-positive patients were less likely to be insured than HCV-negative individuals (61.2% versus 81.2%). Of all HCV-positive patients, 66.7% selleck chemicals were eligible for anti-HCV treatment, but only 54.3% of HCV-positive treatment candidates had any type of insurance coverage, and only 36.3% of treatment-eligible patients had health insurance.27 In contrast, approximately 99% of all Austrians are insured (including jobless people), and treatment for chronic hepatitis B and C is fully covered. A similar insurance coverage rate was reported from Taiwan.28 Thus, there is no direct financial benefit for patients to participate in a study, and no patient contacted our center for the sole purpose of being enrolled in a clinical trial. All patients were referred from general practitioners for evaluation of CHC and were offered treatment, if there were no contraindications. Our center treats approximately one third of all HCV patients from Vienna as well as the surrounding suburbs. These represent the “typical mix” of patients with CHC (36% with history of drug abuse, 19% infected by blood or blood products, and 42% of unknown etiology).

This is also true for the recently licensed protease

inhibitors, which ICG-001 clinical trial markedly improved SVR rates in patients with CHC, GT-1.9-14 As far as we know, this is the first study to assess a potential selection bias in patients with CHC treated within randomized, controlled studies. At least, the outcome of cohort studies in hepatitis C may be influenced by a selection bias.21 Not surprisingly, some baseline characteristics, adherence to treatment, and the IL18B phenotype had the biggest effect on treatment outcome. Patients receiving SOC had more advanced liver disease, a more frequent history of psychiatric disorders, and a substantial proportion was on drug-substitution therapy. Differences in baseline characteristics22-25 reflect inclusion and exclusion criteria and may create “perfect to treat” patients. Whether the slight differences documented in this study are clinically relevant remains unknown, but history of psychiatric disorders, need of drug-substitution therapy, and a higher grade of liver fibrosis may influence treatment outcome, as well. Because of major differences in access to

selleck chemicals llc medical care, data from a European study center cannot be directly applied to the U.S. population of HCV patients. It may also be difficult to draw conclusions from the largely homogenous population of HCV patients seen in Vienna to the largely heterogeneous population of HCV patients seen in the United States. In particular, in this study, except for 7 patients (2.3%; 5 Asians and 2 black Africans), all patients were Caucasians, whereas in the United States, approximately 19% and 40% of treatment eligible or noneligible

patients, respectively, are black.26 Analysis of data from the National Health and Nutrition Examination Survey, conducted in 2005-2008, revealed that HCV-positive patients were less likely to be insured than HCV-negative individuals (61.2% versus 81.2%). Of all HCV-positive patients, 66.7% learn more were eligible for anti-HCV treatment, but only 54.3% of HCV-positive treatment candidates had any type of insurance coverage, and only 36.3% of treatment-eligible patients had health insurance.27 In contrast, approximately 99% of all Austrians are insured (including jobless people), and treatment for chronic hepatitis B and C is fully covered. A similar insurance coverage rate was reported from Taiwan.28 Thus, there is no direct financial benefit for patients to participate in a study, and no patient contacted our center for the sole purpose of being enrolled in a clinical trial. All patients were referred from general practitioners for evaluation of CHC and were offered treatment, if there were no contraindications. Our center treats approximately one third of all HCV patients from Vienna as well as the surrounding suburbs. These represent the “typical mix” of patients with CHC (36% with history of drug abuse, 19% infected by blood or blood products, and 42% of unknown etiology).

7, 34 Identification and consideration of necessary environmental risk factors increase the chances to detect genetic risk factors in association studies.

Biologic interactions between several factors determine the risk of idiosyncratic DILI, and high predictive power will therefore only be achieved if several interacting risk factors are identified in one study,29, 30 and if predictive models can include all genetic as well as environmental risk factors selleck with major contributions. Until recently, genetic risk factors for DILI were mainly studied in case-control candidate gene association studies (CGAS). Meanwhile, high-throughput sequencing technologies with increased speed and lower costs and new methods for the analysis of large complex genetic data sets35, 36 enabled the conduct of large GWAS,37 and the first GWAS have now also been conducted to explore genetics of DILI.14, 38 This section focuses on methodological considerations for the study of DILI in selleck chemicals CGAS and GWAS. General methodological aspects, advantages, and limitations of CGAS and GWAS have been reviewed in detail elsewhere.39, 40 Although CGAS have been able to identify several genetic risk factors for DILI, which are further discussed

below, they have important limitations. First, the selection of candidate genes is guided by current mechanistic knowledge of DILI, and genetic variants that relate to unknown mechanisms may therefore not be detected with an a priori focused search strategy. Second, many CGAS relied on the analysis of a limited number of single-nucleotide polymorphisms and did not consider regions that regulate gene expression;

and even if they targeted the right gene they may have failed to detect a genetic risk factor if the chosen single-nucleotide click here polymorphisms themselves are not relevant for DILI or not in linkage disequilibrium with risk variants. GWAS have the advantage that they are particularly suited for the simultaneous identification of several common low-risk variants in one study. This is relevant for complex diseases like DILI, where the concerted action of several low-risk factors contributes to disease. A priori, GWAS are hypothesis-free and expected to produce a high proportion of false positive results. However, currently used strategies for the identification of genetic risk factors in GWAS extend far beyond simple one-step testing of associations.40 The first GWAS that searched for genetic risk factors of DILI associated with ximelagatran and flucloxacillin successfully used such an extended design with confirmatory analyses in a second set of cases and controls, plus functional in vitro studies.14, 38 These studies also imply that the design not only of CGAS but also of extended multistep GWAS requires guidance by a thorough mechanistic understanding of DILI. This principle is also nicely demonstrated in the introduction and design of two CGAS of DILI associated with diclofenac.

62 The source of liver fat may be adipose tissue because the activation of CB1 receptors in adipocytes promotes lipogenesis,71 and the released fatty HM781-36B acids may be taken up and converted to triglycerides (TGs) by the liver.4 On the other hand, the rapid depletion of excess hepatic TGs after CB1 blockade may involve hepatic CB1 receptors, as indicated by the increased rate of secretion of TG-rich very low density lipoprotein (VLDL) from the livers of both DIO and ob/ob mice after treatment with a peripherally restricted CB1 antagonist62 (see Fig. 2). Endocannabinoids are also involved in the

diet-induced decrease in fatty acid oxidation. The activity of hepatic carnitine palmitoyltransferase 1 (CPT1), the rate-limiting enzyme in mitochondrial fatty acid β-oxidation, is suppressed by either a high-fat diet or treatment with a

CB1 agonist, and both effects are prevented by rimonabant.24 Conversely, hepatic CPT1 activity is increased in CB1−/− mice24 and in DIO mice after chronic CB1 blockade.24, 62, 72 Adiponectin is a key stimulator of fatty acid β-oxidation, and CB1 blockade increases plasma adiponectin.73 The improved insulin sensitivity following CB1 blockade Navitoclax price has been found to have both adiponectin-dependent74, 75 and adiponectin-independent components,75 although the role of adiponectin in the effects of CB1 blockade on hepatic mitochondrial function and fatty acid oxidation has not been explored. Increased energy expenditure due to increased fat oxidation after CB1 blockade has been documented with indirect selleck chemicals llc calorimetry in rats76-78 and mice.62 These effects likely contribute to the food intake–independent sustained weight loss62, 79 as well

as the reversal of hepatic steatosis62, 80, 81 after chronic CB1 blockade. The DIO-related hypertriglyceridemia was modestly attenuated, whereas the accompanying increase in plasma LDL cholesterol and decrease in high-density lipoprotein cholesterol were absent in both CB1−/− and LCB1−/− mice on a high-fat diet. This suggests that hepatic CB1 mediates diet-induced changes in hepatic lipoprotein metabolism and/or secretion. In a recent study, the treatment of mice with an inhibitor of monoglyceride lipase resulted in elevated hepatic levels of 2-AG, increased hepatic expression of sterol regulatory element binding protein 1c (SREBP1c) and FAS, hypertriglyceridemia, and an accumulation in plasma of apolipoprotein E (ApoE)–depleted, TG-rich apolipoproteins.68 These changes were absent in CB1−/− and ApoE−/− mice and could be prevented by CB1 blockade. Furthermore, despite the elevated hepatic lipogenic gene expression, TG secretion rates were unchanged, but TG clearance from plasma was inhibited.

62 The source of liver fat may be adipose tissue because the activation of CB1 receptors in adipocytes promotes lipogenesis,71 and the released fatty ABT-263 purchase acids may be taken up and converted to triglycerides (TGs) by the liver.4 On the other hand, the rapid depletion of excess hepatic TGs after CB1 blockade may involve hepatic CB1 receptors, as indicated by the increased rate of secretion of TG-rich very low density lipoprotein (VLDL) from the livers of both DIO and ob/ob mice after treatment with a peripherally restricted CB1 antagonist62 (see Fig. 2). Endocannabinoids are also involved in the

diet-induced decrease in fatty acid oxidation. The activity of hepatic carnitine palmitoyltransferase 1 (CPT1), the rate-limiting enzyme in mitochondrial fatty acid β-oxidation, is suppressed by either a high-fat diet or treatment with a

CB1 agonist, and both effects are prevented by rimonabant.24 Conversely, hepatic CPT1 activity is increased in CB1−/− mice24 and in DIO mice after chronic CB1 blockade.24, 62, 72 Adiponectin is a key stimulator of fatty acid β-oxidation, and CB1 blockade increases plasma adiponectin.73 The improved insulin sensitivity following CB1 blockade Belinostat mw has been found to have both adiponectin-dependent74, 75 and adiponectin-independent components,75 although the role of adiponectin in the effects of CB1 blockade on hepatic mitochondrial function and fatty acid oxidation has not been explored. Increased energy expenditure due to increased fat oxidation after CB1 blockade has been documented with indirect click here calorimetry in rats76-78 and mice.62 These effects likely contribute to the food intake–independent sustained weight loss62, 79 as well

as the reversal of hepatic steatosis62, 80, 81 after chronic CB1 blockade. The DIO-related hypertriglyceridemia was modestly attenuated, whereas the accompanying increase in plasma LDL cholesterol and decrease in high-density lipoprotein cholesterol were absent in both CB1−/− and LCB1−/− mice on a high-fat diet. This suggests that hepatic CB1 mediates diet-induced changes in hepatic lipoprotein metabolism and/or secretion. In a recent study, the treatment of mice with an inhibitor of monoglyceride lipase resulted in elevated hepatic levels of 2-AG, increased hepatic expression of sterol regulatory element binding protein 1c (SREBP1c) and FAS, hypertriglyceridemia, and an accumulation in plasma of apolipoprotein E (ApoE)–depleted, TG-rich apolipoproteins.68 These changes were absent in CB1−/− and ApoE−/− mice and could be prevented by CB1 blockade. Furthermore, despite the elevated hepatic lipogenic gene expression, TG secretion rates were unchanged, but TG clearance from plasma was inhibited.

The Declaration of Helsinki Principles was followed, and all participants gave written consent for participation in this study. The studies in animals were conducted in accordance with European Union

guidelines (86/609/CEE) and French National Chart guidelines, and protocols were approved by the local Ethics Committee for Animal Experimentation of Grenoble (ComEth). Ninety-four HLA-A*0201+ chronic HBV-infected patients and one resolved control were studied. HBV patients (Table 1) were classified as inactive carriers (HBeAg-negative, HBV-DNA <2,000 IU/mL, and consistently normal alanine aminotransferase [ALT] for at least 1 year), HBeAg-negative hepatitis, and HBeAg-positive hepatitis. Forty-eight patients were treated

(entecavir/tenofovir), and HBV-DNA was undetectable in 83% of these patients. Exclusion criteria included Ensartinib supplier human immunodeficiency virus/hepatitis C virus/hepatitis D virus coinfection, other liver diseases, and treatment with IFN-α or immunosuppressive agents. Serum HBs antigen was quantified using the Abbott Architect HBsAg QT assay Panobinostat mouse (Abbott Diagnostics). Samples were also obtained from HLA-A*0201+ healthy donors. PBMCs were purified via Ficoll-Hypaque density-gradient centrifugation (Eurobio). Liver tissues, obtained from six HLA-A*0201+ HBV patients (Table 1), were processed to prepare liver-infiltrating lymphocytes (LILs). From all liver biopsies, we obtained 0.45 × 106 to 2.6 × 106 cells, among which 14.2%-58.2% were CD3+ T cells and 8.3%-43.3% were CD8+ T cells. The GEN2.2 pDC line was cultured as described.26 The HLA-A*0201+ hepatocyte line HepG2 was cultured in Dulbecco’s modified Eagle’s medium, 10% selleckchem fetal bovine serum, 50UI/ml penicillin/streptomycin (Invitrogen), 1 mM sodium pyruvate (Sigma). HepG22.15 (HepG2 transfected with HBV-DNA) was cultured in William’s E, 10% fetal

bovine serum, 50 IU/mL penicillin/streptomycin, 2 mM Glutamine (Invitrogen), 5 μg/mL insulin (Sigma) and 5.10−5 hydrocortisone hemisuccinate (Roche). All other cultures were performed in RPMI1640-Glutamax, 1% nonessential amino acids, 100 μg/mL gentamycin, 10% fetal bovine serum (Invitrogen), and 1 mM sodium pyruvate (Sigma). T2 and K562 lines were purchased from American Type Culture Collection (LGC Standards). We used the following HLA-A*0201-restricted peptides (NeoMPS) and corresponding HLA-A*0201 tetramers (Beckman): HBc18-27 (FLPSDFFPSV; core), HBs335-343 (WLSLLVPFV; surface), pol575-583 (FLLSLGIHL; polymerase), and FluM158-66 (GILGFVFTL; influenza matrix). The pDC line was loaded with peptides as described.26 PBMCs or LILs were cocultured with peptide-loaded pDCs at a 1:10 ratio and restimulated weekly in presence of 200 IU/mL IL-2 (Proleukine, Chiron). In some experiments, PBMCs were directly stimulated with the peptide (1-10 μM final) for 10 days.