Search

The purpose of this study was to monitor abundance and activation of local CD8β-expressing T-cell populations during Eimeria tenella infections of naïve chickens and chickens immune by previous infections. Chickens were infected with E. tenella up to three times. Caecal T-cell receptor ( TCR) γ/δ- CD8β+ cells (cytotoxic T lymphocytes; CTL) and TCRγ/δ+ CD8β+ cells were characterized with respect to activation markers (blast transformation, CD25 and cell surface CD107a). Cells were also induced to degranulate in vitro as a measure of activation potential. Major findings included a prominent long-lasting, up to 6 weeks, increase in the proportion of CTL among caecal CD45+ cells in the later stages after primary E. tenella infection. These CTL also showed clear signs of activation, that is blast transformation and increased in vitro induced degranulation. At second and third E. tenella infection, chickens showed strong protective immunity but discrete signs of cellular activation were observed, for example increased in vitro induced degranulation of CTL. Thus, primary E. tenella infection induced clear recruitment and activation of local CTL. Upon subsequent infections of strongly immune chickens cellular changes were less prominent, possibly due to lower overall numbers of cells being activated because of the severe restriction of parasite replication. [ABSTRACT FROM AUTHOR]

Abstract: CD4 and CD8 molecules are co-receptors of T cell receptors which interact specifically with MHC class II and I, respectively, during antigen presentation. Here we investigated CD4 and CD8 expression patterns in a fish, Japanese flounder, Paralichthys olivaceus in response to infection and tuberculin injection. The CD4-1 mRNA level was gradually and weakly increased in trunk kidney after infection with Streptococcus iniae, Edwardsiella tarda and viral hemorrhagic septicemia virus (VHSV), while the CD4-2 mRNA level was dramatically increased after E. tarda and VHSV infection, but not increased after S. iniae infection. CD4-2 mRNA but not CD4-1mRNA increased in the kidney during tuberculin response which is mediated by memory Th1 cells. The patterns for the change of mRNA level in CD8α and CD8β were similar to those of the CD4-2 during the infections and tuberculin response. Fluorescent in situ hybridization detected CD4-1 mRNAs on melano-macrophage centers and CD4-2 mRNAs at some cell clusters located near the melano-macrophage centers. CD8α and CD8β mRNAs were detected at the same cell clusters in the spleen and head kidney. These results suggest that CD4-1 and CD4-2 are expressed in different cells and that CD4-2-positive cells, rather than CD4-1-positive cells, have a main role in Th1-related immune responses collaborating with CD8α- and CD8β-positive cells in Japanese flounder. [Copyright &y& Elsevier]

The full-length complementary DNA (cDNA) sequences encoding cd8α and cd8β molecules were sequenced and characterized from mandarin fish Siniperca chuatsi. Conserved motifs and residues were found to be present in derived peptides of the Cd8 molecules. For example, WXR motif, DXGXYXC motif, and four cysteine residues were present in the extracellular region of the Cd8 protein. Threonine, serine and proline residues involved in multiple O-linked glycosylation events were located in the membrane proximal hinge region. The common CPH motif in the cytoplasmic tail was detected similar to other teleost Cd8 molecules. Different from those in mammals, S. chuatsi Cd8 sequences have many extra cysteine residues (C149 in Cd8 α sequence and C46, C51 and C158 in Cd8 β sequence), which also exist in other teleost Cd8 molecules. Real-time polymerase chain reaction (RT-PCR) and Western blot analyses revealed that the thymus had the highest expression of cd8 messenger (m)RNA and protein. After stimulated with phytohaemagglutinin, polyriboinsine-polyribocyaidylic acid and concanavalin A (ConA), the expression level of cd8 mRNA increased significantly in head-kidney lymphocytes at 4 and 8 h, but decreased to normal level at 12 h. Similarly, stimulation with ConA in vivo also led to an increase in the cd8 mRNA level in the spleen. Immunohistochemistry analysis demonstrated that Cd8 α-positive cells can be detected in the thymus, spleen and intestine by using polyclonal anti-Cd8 α antibody. [ABSTRACT FROM AUTHOR]

Abstract: The transcript levels of viral RNAs, selected T-cell marker and cytokine genes, toll like receptor (TLR) 7, and two interferon stimulated genes (ISG) were analysed in sexually immature adult Atlantic halibut (Hippoglossus hippoglossus L.) experimentally infected with nodavirus. The expression of the T-cell markers, TLR7 and the cytokine genes was further explored in in vitro stimulated anterior kidney leucocytes (AK leucocytes) isolated from the experiment fish and from additional untreated non-injected fish. The levels of viral RNA1 and RNA2 were increasing in brain and eye at around 4 and 8weeks post injection (wpi), respectively, and still increasing at the end of the experiment, especially in eye. Immuno-positive cells and signs of vacuolisation in both brain and eye were seen at 14wpi. Increased transcript levels of TCRβ, CD4-2, CD4, CD8α, and Lck in brain and eye of the experimentally infected halibut suggested an involvement of halibut T-cells in the immune response against nodavirus. Interestingly, a similar expression pattern of TCRβ, CD4 and Lck was seen in both brain and eye. However, compared to brain that showed elevated transcript levels of TCRβ, CD4 and Lck mainly at 10 and 14wpi, the increase appeared earlier between 3 and 4wpi in the eye. Yet, an increase in the transcript level of IFNγ was seen at 10 and 14wpi in both organs. Moreover, elevated levels of TLR7, IL-1β, IL-6, ISG15 and Mx were detected in vivo. The in vitro experiments, stimulating AK leucocytes with ConA–PMA, imiquimod or nodavirus, further supported an involvement of IL-6 and IFNγ in the immune response against nodavirus and the involvement of CD8β+ cells. Results from the present study thus indicate an importance of T-cells, IFNγ and the analysed ISGs in the immune response against nodavirus in Atlantic halibut, and would be of great help in future vaccination trials giving the possibility to monitor the immune response rather than mortality during post-vaccination challenge experiments. [Copyright &y& Elsevier]

Abstract: The cytoplasmic tail of mammalian CD8α binds the kinase LCK in a zinc-dependent manner. In analogy with a previous study for humans (Kim et al., 2003) peptides were synthesized from rainbow trout CD8α and LCK. Surface plasmon resonance (SPR) analysis indicated that also in fish these molecules bind to each other in a zinc-dependent manner. [Copyright &y& Elsevier]

Abstract: Partial cDNA sequences of both CD8β and CD4-like (CD4L) genes of common carp (Cyprinus carpio L.) were isolated from thymus cDNA library by the method of suppression subtractive hybridization (SSH). Subsequently the full length cDNAs of carp CD8β and CD4L were obtained by means of 3′ RACE and 5′ RACE, respectively. The full length cDNA of carp CD8β is 1164bp and encodes 207 amino acids including a signal peptide region of 24 amino acids, a transmembrane region of 23 amino acids from aa 167 to aa189 and an immunoglobulin V-set from aa 19 to aa 141. Similar to other species CD8βs, carp CD8β also lacks p56lck domain in the cytoplasmic region. The full length cDNA of carp CD4L is 2001bp and encodes 458 amino acids including four immunoglobulin (Ig)-like domains in the extracellular region, a transmembrane region of 23 amino acids at the C-terminal region from aa 402 to aa 424 and a cytoplasmic tail. Similar to mammalian, avian CD4s and fugu CD4L, carp CD4L also has the conserved p56lck tyrosine kinase motif (C-X-C) in the cytoplasmic region. RT-PCR analysis demonstrated that carp CD8β and CD4L genes were both expressed predominantly in thymus. The results from this study can be used to understand the evolution of both the CD8β and CD4 molecules which can be used as markers for cytotoxic and helper T cells in carp. [Copyright &y& Elsevier]

Interleukln (IL-4) can induce CD8 expression on mature CD4 T cells. To study this phenomenon in more detail, we characterized CDS expressed on IL-4-lnduced CD4 CD8 (double positive) T cell clones in comparison with that on CD8 T cell clones. Using 2ST8–5H7 mAb that detects CD8β expression, we found that double positive T cell clones isolated with IL-4 express CD8α but not β, in contrast to CD8 CTL cell clones, which express both chains of CD8. Northern blot analysis revealed that these double positive clones expressed CD8α but not β mRNA, indicating that CD8α and β are Independently regulated at the pre-translatlonal level. Immunopreclpitation experiments showed that CD8 expressed on a representative IL-4-lnduced double positive T cell clone consists mainly of homodimers of a single 34 kd protein of CD8α. The amount of multimers detected from this clone was much less than that from a CD8+ CTL clone. These results suggest that persistent expression of CD8β Is specific for the CD8+ lineage and may be involved in polymerization and stabilization of CD8 which enhances the efficiency of class l-restrlcted antigen recognition. [ABSTRACT FROM PUBLISHER]

Expression and function of mouse and human CD8 (mCD8 and hCD8) α chain molecules In mouse T cell hybridomas were analyzed. The expression of cytolytic T lymphocyte-derived CD8 molecules was supprmsd In hybridomas established by fusing the BW5147 thymoma to a CD8 cytolytic T lymphocyte clone, while expression of CD4 remained intact In BW × CD4 helper T cell hybridomas. However, hybridomas established by tusing a cytolytlc T cell clone wtth BW5147 cell lines, transfected wlth either the mCD8 or hCD8 chaln, expressed the T cell-derived mCD8 chain as a CD8 heterodimer (mCD8/mCD8 or hCD8/mCD8). These data suggest that negative regulatory mechanlsms for the mCD8 gene In BW thymoma failed to suppress mCD8 gene expression, indicating different regulatory mechanisms for the tightly linked mCDh and mCD8 genes. Analysis of the antlgen reactivity of the hybridomas revealed that the human CDB chain felled to increase the mouse T cell receptor - class I MHC Interaction, even as a heterodimeric form with mCD88 molecules. However, both the human mCD8 homodimer and the heterodimeric form with mCD88 were found to be capable of suppressing the class 11-restricted T cell response. [ABSTRACT FROM PUBLISHER]