3B) This suggests that T lymphocytes from the nonresponder group

3B). This suggests that T lymphocytes from the nonresponder group are generally compromised in their ability to respond to a specific antigen following major histocompatibility complex–dependent presentation by an antigen-presenting cell. No significant

correlation was found between the inability to respond to the HBc-loaded pDC stimulation and HBV-DNA levels (Fig. 3C), HBs antigen level (Fig. 3D), Selleckchem Mitomycin C ALT measurements (Fig. 3E), or antiviral treatment (Fig. 3F). In contrast, the presence of HBeAg in the serum appeared to differentiate between responder and nonresponder chronic HBV patients (Fig. 4). The HBc-specific T cell response was much greater in inactive carriers and treated or untreated HBeAg-negative hepatitis BIBW2992 cost patients than in HBeAg-positive patients (Fig. 4A). After pooling patients according to HBeAg status alone, this difference appeared clearly significant (Fig. 4B). This interesting observation was corroborated by data for two patients in whom HBeAg status changed over a

6-month interval (Fig. 4C,D). One HBeAg-positive patient, unexpectedly capable of responding to pDC stimulation, achieved HBeAg loss followed by HBeAg seroconversion 6 months later. The other patient, initially HBeAg-negative and capable of responding to HBc-loaded pDC stimulation, became unresponsive 6 months later during a transient HBeAg-positive peak. Thus, HBeAg status distinguishes between chronic HBV patients capable of responding, or not, to HBc-loaded pDC

stimulation. To investigate the functionality of HBV-specific T cells generated from responder chronic HBV patients we examined T cell exhaustion and cytotoxic potential. PD1 expression, a marker of T cell exhaustion, was not detected on the HBc-specific T cells elicited by the pDC line (Supporting Fig. 1). The cytotoxic potential of expanded HBV-specific T cells was determined by performing a 51Cr release assay using peptide-loaded HLA-A*0201+ T2 cells as targets. As expected, HBc-specific T cells exhibited a strong cytotoxicity toward T2 cells loaded with HBc peptide but not with an irrelevant peptide, showing the specificity of the HBV-specific T cells function (Fig. 5A). Next, we tested the ability of these specific T cells to lyse a more relevant target, such MG-132 ic50 as HBV-transfected HLA-A*0201+ hepatocytes. Due to the lack of P3 facilities necessary to perform radioactive experiments with virus-producing cells, a CFSE assay was used. This assay consisted of culturing specific T cells with a mixture of two targets labeled with distinct CFSE intensities. The disappearance of the CFSE pic, as measured via flow cytometry, indicates killing of the corresponding cells. For all patients tested, HBc-specific T cells were able to specifically lyse the HBV-transfected HLA-A*0201+ hepatocyte cell line HepG22.15, but not the HBV-free HepG2 line (Fig. 5B).

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