HIV-1 Infection Abrogates CD8<sup>+</sup>T Cell Mitogen-Activated Protein Kinase Signaling Responses

Crawford, Timothy Q.; Ndhlovu, Lishomwa C.; Tan, Alice J.; Carvidi, Alex; Hecht, Frederick M.; Sinclair, Elizabeth; Barbour, Jason D.

doi:10.1128/jvi.05682-11

Cited by 24 publications

(34 citation statements)

References 31 publications

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“…Serum HBsAg and HBV DNA were monitored at day 7 after the second and third immunization. The inhibition rate of serum HBsAg or viral DNA in HBV transgenic mice of CTPHBcAg 18-27 -Tapasin group was higher than CTP-HBcAg [18][19][20][21][22][23][24][25][26][27] or HBcAg 18-27 -Tapasin or IFN-a (Figures 4a and b). Serum HBsAg level from the mice immunized with CTP-HBcAg 18-27 -Tapasin decreased markedly as compared with the mice immunized with CTP-HBcAg 18-27 , HBcAg 18-27 -Tapasin, or IFN-a (*Po0.05).…”

Section: Discussionmentioning

confidence: 99%

“…Phorbol 12-myristate 13-acetate (PMA), ionomycin, monensin, phytohemagglutinin (PHA), and concanavalin A (ConA) were obtained from Sigma (USA). Soluble fusion proteins CTP-HBcAg 18-27 -Tapasin, CTP-HBcAg [18][19][20][21][22][23][24][25][26][27] , and HBcAg 18-27 -Tapasin were purified and had undetectable endotoxin level according to our former experiments (Figures 1a and b). 19 EL-4 cells (H-2K b ; from ATCC) were cultured in Dulbecco's modified Eagle's medium containing 10% FBS at 37 1C under a humidified condition of 5% CO 2 and 100 U/ml penicillin.…”

Section: Materials and Methods Reagents Cells And Fusion Proteinsmentioning

confidence: 99%

“…Splenocytes were stimulated in the presence of 10 mg/ml HBcAg for 6 h. After incubation for 3 h, 25 mg/ml of PMA, 1 mg/ml of ionomycin, and 1.7 mg/ml of monensin were added and incubation continued for another 3 h. 23 After washing with PBS, cells were stained with saturating concentrations of PE-conjugated anti-CD8a McAb. After being fixed with Fix and Perm reagents A and B (BD Biosciences, USA), the cells were incubated for 20 min with FITC-labeled anti-IFN-g McAb, followed by washing two times with PBS and analyzing by flow cytometry.…”

Section: Measurement Of Ifn-g Production By Intracellular Cytokine Stmentioning

confidence: 99%

“…41,42 HBcAg 18-27 epitope has been shown to be able to induce HBV-specific CTL response and thus is a potential for the therapeutic design of chronic HBV infection. 16,17 In the present study, we expressed HBcAg [18][19][20][21][22][23][24][25][26][27] included the complete amino-acid sequences that were fused with CTP and Tapasin. Meanwhile, our previous results showed that recombinant CTP-HBcAg 18-27 -Tapasin could penetrate into CTP-HBcAg 18-27 -Tapasin induces HBV-specific CTLs X Chen et al DCs cytoplasm.…”

Section: Ctp-hbcag 18-27 -Tapasin Induces Hbv-specific Ctls X Chen Et Almentioning

confidence: 99%

“…12,13 The low surface expression of MHC I molecules resulted in impaired positive and negative thymic T-cell selection, resulting in a reduction of CD8 þ T-cell number in the thymus and periphery, and reduced antigen presentation to T cells, resulting in poor in vivo CTL response to influenza virus. 14,15 HBcAg 18-27 epitope (HBcAg [18][19][20][21][22][23][24][25][26][27] ) is a widely accepted as a CTL epitope to stimulate CTL response to hepatitis B core antigen (HBcAg). 16,17 However, HBV-specific CTL responses induced by exogenous antigens such as HBV-encoded antigens are usually weak, mainly because of the prohibition of intracellular delivery of antigens and peptides by the lipophilic nature and selective permeability of biological membranes.…”

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Tapasin modification on the intracellular epitope HBcAg18–27 enhances HBV-specific CTL immune response and inhibits hepatitis B virus replication in vivo

Chen

Tang

Zhang

et al. 2014

Laboratory Investigation

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HBV-specific cytotoxic T-lymphocyte (CTL) activity has a very important role in hepatitis B virus clearance. Present studies suggest that Tapasin, a endoplasmic reticulum (ER) chaperone, stabilizes the peptide-receptive MHC I conformation, allowing peptide exchange and increasing more peptides to be translocated into the ER. We have previously testified that cytoplasmic transduction peptide (CTP)-HBcAg 18-27 -Tapasin fusion protein could enter cytoplasm of dendritic cells, and enhance T cells' response to generate specific CTLs efficiently in vitro. In the present study, we evaluated specific immune responses of CTP-HBcAg 18-27 -Tapasin fusion protein in HLA-A2 transgenic mice (H-2K b ) and anti-viral ability in HBV transgenic mice, and explored the mechanisms probably involved in. The studies showed that CTP-HBcAg 18-27 -Tapasin not only increased production of cytokine IFN-g and interleukin-2 (IL-2), compared with CTP-HBcAg 18-27 , HBcAg 18-27 -Tapasin, and PBS, but also significantly induced the higher percentages of IFN-g þ CD8 þ T cells and specific CTL responses in HLA-A2 transgenic mice. Moreover, enhancement of specific CTL activity induced by the fusion protein reduced HBV DNA and hepatitis B surface antigen (HBsAg) levels and decreased the expression of HBsAg and hepatitis B core antigen (HBcAg) in liver tissue of HBV transgenic mice. In addition, CTP-HBcAg 18-27 -Tapasin could upregulate the expression of JAK2, Tyk2, STAT1, and STAT4 in T lymphocytes in HLA-A2 transgenic mice splenocytes. However, there was no significant difference on the expressions of JAK1, JAK3, and STAT6 between each group. In conclusion, CTP-HBcAg 18-27 -Tapasin fusion protein could enhance not only the percentages of CTLs but also induce robust specific CTL activity and inhibits hepatitis B virus replication in vivo, which was associated with activation of the JAK/STAT signaling pathway. Hepatitis B virus (HBV) infection remains a global problem, despite the effectiveness of the HBV vaccines in preventing infection. Although the factors that contribute to the clearance of the virus in acute self-limiting HBV infection or to the persistence of the virus in chronic HBV infection are not completely clear, increasing evidence suggests that host immune responses are an important factor in determining the outcome of HBV infection. 1,2 Acute individuals develop a strong, polyclonal, multispecific cytotoxic T-lymphocyte (CTL) response and a polyclonal T helper (Th) cell response to the virus typically, while these responses are markedly attenuated in chronically infected patients. 3,4 Persistent HBV infection is characterized by a weak adaptive immune response, thought to be due to inefficient CD4 þ T-cell priming in the early stage of virus infection and subsequent development of a quantitatively and qualitatively ineffective CD8 þ T-cell response. Studies of HBV infection in woodchucks and chimpanzees, as well as patients, are suggesting that multiple and frequent administration of the vaccine may be advisable in treatment of chronic...

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Section: Discussionmentioning

confidence: 99%

Section: Materials and Methods Reagents Cells And Fusion Proteinsmentioning

confidence: 99%

Section: Measurement Of Ifn-g Production By Intracellular Cytokine Stmentioning

confidence: 99%

Section: Ctp-hbcag 18-27 -Tapasin Induces Hbv-specific Ctls X Chen Et Almentioning

confidence: 99%

mentioning

confidence: 99%

See 3 more Smart Citations

Tapasin modification on the intracellular epitope HBcAg18–27 enhances HBV-specific CTL immune response and inhibits hepatitis B virus replication in vivo

Chen

Tang

Zhang

et al. 2014

Laboratory Investigation

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Concomitant evaluation of PMA+ionomycin‐induced kinase phosphorylation and cytokine production in T cell subsets by flow cytometry

et al. 2014

Self Cite

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Methods to detect intracellular kinase signaling intermediates by flow cytometry have been recently developed. Termed "phospho-flow," these methods employ fluorescenceconjugated monoclonal antibodies that recognize phosphorylated epitopes of intracellular kinases, and may be combined with surface phenotypic markers to observe changes in kinase pathways by cellular subset. Effector functions, like cytokine production, are processes intrinsically linked to intracellular signaling and kinase activity within each cell. Methodologies that would simultaneously detect changes to signaling pathways as well as effector responses at the single-cell level would allow for mapping of the functional consequences induced by signaling pathway modifications. However, there are challenges to developing such a combined protocol, relating to the different kinetics of rapid signaling events and the more prolonged time required to induce and observe cytokine responses. In this report, we describe the development of an assay that accommodates differences in protocol conditions and response kinetics, merging phospho-flow cytometry, and intracellular cytokine staining methods into a single experimental protocol. We examined intracellular ERK1/2 phosphorylation and IFN-c production by CD41 and CD81 T cells upon polyclonal stimulation with PMA and ionomycin, while monitoring expression of the cytolytic molecule perforin and the T cell activation marker CD38. We present a method that allows observation of kinase phosphorylation and cytokine production within the same cell after stimuli, while maintaining a stable cellular phenotype. Monitoring of signaling and effector functions in distinct immune subsets provides a platform to investigate and relate intracellular kinase signaling activity to immune cell effector function and phenotype in disease states. V C 2014 International Society for Advancement of Cytometry Key terms T cell; ERK; IFN-c; perforin; CD38; HLA-DR; phosflow; phospho-flow intracellular cytokine staining; immune effector function; ICS Flow cytometry technology allows for detection of cellular functional responses, such as the production of inflammatory cytokines using intracellular cytokine staining (ICS) (1-4), or more recently, the phosphorylation of intracellular kinase signaling intermediates using phospho-specific antibodies (phospho-flow) (5-10). Evaluation of kinase phosphorylation and cytokine production simultaneously in a singleflow cytometry experiment allows examination of associations between signaling pathway activity and effector functions. This approach will be of interest in human diseases, such as autoimmune conditions, viral infections, and cancers, where changes to cellular kinase activity have effects on immune cell function and contribute to disease pathogenesis (11-13).We previously observed an abrogation of extracellular signal-regulated kinase (ERK1/2) signaling in subpopulations of activated CD81 T cells in adults during early

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CD38 is a putative functional marker for side population cells in human nasopharyngeal carcinoma cell lines

Zheng

Liao

Zhu

et al. 2015

Molecular Carcinogenesis

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Cancer stem cells (CSCs) are thought to be responsible for cancer progression and therapeutic resistance but identification of this subpopulation requires selective markers. Fortunately, side population (SP) cells analysis brings a novel method to CSCs study. In this study, we identified SP cells, which are demonstrated rich in CSCs, in four nasopharyngeal carcinoma (NPC) cell lines. We investigated SP cells from HK-1 NPC cell line and showed CSCs characteristics in this subpopulation. SP cells displayed greater proliferation and invasion and expressed high levels of CSCs markers than NSP cells. Furthermore, our microRNA microarray analysis of SP versus NSP cells revealed that CD38-related miRNAs were down-regulated in SP cell, but the mRNA and protein level of CD38 were highly expressed in SP cells. We further searched for molecules interacting with CD38 and identified ZAP70, which was also well expressed in SP cells at both mRNA and protein levels. Our results uncover a CD38 pathway that may regulate the proliferation and migration of SP cells from HK-1 NPC cell line.

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HIV-1 Infection Abrogates CD8⁺T Cell Mitogen-Activated Protein Kinase Signaling Responses

Cited by 24 publications

References 31 publications

Tapasin modification on the intracellular epitope HBcAg18–27 enhances HBV-specific CTL immune response and inhibits hepatitis B virus replication in vivo

Tapasin modification on the intracellular epitope HBcAg18–27 enhances HBV-specific CTL immune response and inhibits hepatitis B virus replication in vivo

Concomitant evaluation of PMA+ionomycin‐induced kinase phosphorylation and cytokine production in T cell subsets by flow cytometry

CD38 is a putative functional marker for side population cells in human nasopharyngeal carcinoma cell lines

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HIV-1 Infection Abrogates CD8+T Cell Mitogen-Activated Protein Kinase Signaling Responses

Cited by 24 publications

References 31 publications

Tapasin modification on the intracellular epitope HBcAg18–27 enhances HBV-specific CTL immune response and inhibits hepatitis B virus replication in vivo

Tapasin modification on the intracellular epitope HBcAg18–27 enhances HBV-specific CTL immune response and inhibits hepatitis B virus replication in vivo

Concomitant evaluation of PMA+ionomycin‐induced kinase phosphorylation and cytokine production in T cell subsets by flow cytometry

CD38 is a putative functional marker for side population cells in human nasopharyngeal carcinoma cell lines

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HIV-1 Infection Abrogates CD8⁺T Cell Mitogen-Activated Protein Kinase Signaling Responses