T-cell Receptor Delta Research Articles

Human TcR gamma delta+ lymphocyte response on primary exposure to Plasmodium falciparum.

In 29 patients experiencing their first P. falciparum malarial attack, blood levels of TcR gamma delta+ lymphocytes were studied from the onset of infection to up to 6-9 months later. Blood TcR gamma delta+ lymphocytes, revealed using the TcR delta 1 monoclonal antibody (MoAb), were increased both in absolute and relative numbers. Alterations lasted for up to 3-4 months following the attack. A Ti gamma A/BB3 reactive V gamma 9 subset was preferentially amplified. In vitro, TcR gamma delta+ lymphocytes from both malaria-sensitized and unprimed donors responded to P. falciparum schizont extract (PFSE). PFSE-stimulated polyclonal T cell lines consisted principally in TcR gamma delta+ cells with a Ti gamma A+/BB3+ phenotype. Several TcR gamma delta+ T cell clones obtained from patients recovering from acute malarial attack were maintained in the presence of PFSE and autologous irradiated PBL. They belong to the V gamma 9 subset. In long-term cultures, TcR gamma delta+ clones progressively lost their capacity to react to PFSE antigen while they were able to proliferate and to exert cytotoxic activity in response to autologous TcR alpha beta+, PFSE-specific T lymphocyte clones. This suggests that regulatory interactions occur between activated TcR gamma delta+ and TcR alpha beta+ cells generated by P. falciparum. Sequential variations in blood TcR gamma delta+ and TcR alpha beta+ lymphocyte levels after primary exposure to P. falciparum suggest that such regulatory interactions may occur in vivo.

Characterization of fresh (uncultured) tumour-infiltrating lymphocytes (TIL) and TIL-derived T cell lines from patients with renal cell carcinoma.

Fresh (uncultured) TIL from 12 untreated patients with primary renal cell carcinoma were prepared from tumour specimens by enzymatic digestion, and were characterized by immunofluorescence using MoAbs recognizing leucocyte differentiation antigens or particular V alpha or V beta segments of the T cell receptor (TCR). These fresh TIL comprised CD3+ (20-84%); CD4+ (3-15%); CD8+ (13-35%); alpha beta TCR+ (20-50%); gamma delta TCR+ (3-17%); CD16+ (1-18%) and CD56+ (3-10%) cells. Significant proportions of V alpha 2+, V beta 5.1+ and V beta 6+ cells were found in TIL of certain patients with renal cell carcinoma, suggesting that they comprised oligoclonal T cells. T cell lines were developed in low concentrations of rIL-2 (200 U/ml) from TIL from 11 patients with renal cell carcinoma, and were characterized by immunofluorescence and cell-mediated cytotoxicity. These T cell lines consisted primarily of CD3+ (51-94%); CD4+ (1-80%); CD8+ (0-84%); alpha beta TCR+ (65-87%); gamma delta TCR+ (0-25%); CD16+ (0-16%) and CD56+ (2-57%) cells. These T cell lines exhibited non-specific cytotoxicity against autologous and allogeneic renal tumour cells, with the exception of one T cell line that exhibited preferential cytotoxicity against autologous renal tumour cells. These results suggest that fresh TIL from patients with renal cell carcinoma contain significant proportions of oligoclonal T cells that may have accumulated at the tumour site as a result of a clonal expansion.

T cell receptor repertoire in the peripheral blood and intestinal mucosa of coeliac patients.

The alpha beta and gamma delta T cell receptor (TCR) repertoire in the peripheral blood and intestinal mucosa of six coeliac and six age-matched controls was analysed by reverse transcription and polymerase chain reaction (PCR). No TCR alpha and gamma delta restriction was observed in coeliacs and controls. However, V gamma 3 was expressed only in coeliac peripheral and intestinal T cells. V delta 2 was strongly expressed in coeliacs and scarcely transcribed in control cells. The unique expression of these gamma delta TCR in coeliac patients suggests that V gamma 3 and perhaps V delta 2 TCR-bearing lymphocytes may play a role in the pathogenesis of coeliac disease.

Gamma delta T cell receptor gene expression by muscle-infiltrating lymphocytes in the idiopathic inflammatory myopathies.

Autoreactive alpha beta T cells have been implicated as playing a primary pathogenic role in a group of diseases characterized by chronic muscle inflammation known as the idiopathic inflammatory myopathies (IIM). gamma delta T cells, a distinct and enigmatic class of T cells, play a less certain role in a variety of human autoimmune diseases including the IIM. In an attempt to understand the significance of gamma delta T cells in the IIM, we utilized a sensitive polymerase chain reaction (PCR) technique to evaluate gamma delta T cell receptor (TCR) gene expression in 45 muscle biopsies obtained from 42 IIM patients (17 polymyositis, 12 dermatomyositis, and 13 inclusion body myositis). gamma delta TCR gene expression was not detected in 36 specimens, the majority of muscle biopsies surveyed. gamma delta TCR gene expression by muscle-infiltrating lymphocytes was detected among nine clinically heterogeneous patients. We further analysed the junctional sequence composition of the V gamma 3 and V delta 1 transcripts, whose expression was prominent among gamma delta positive patients. DNA sequence analysis of V gamma 3 amplification products from two patients revealed the presence of several productively rearranged transcripts with amino acid sequence similarities within the V gamma 3-N-J gamma junctional domain. No amino acid sequence similarities were evident within the V delta-N-D delta-N-J delta region of V delta 1 transcripts amplified from four patients, although a distinct and dominant clonotype was detected from each patient. Our cumulative data suggest that unlike alpha beta T cells, gamma delta T cells do not play a prominent pathologic role in the IIM. In fact, the sporadic nature of gamma delta TCR gene expression detected among these patients implies that gamma delta T cell infiltration, when it occurs, is a secondary event perhaps resulting from non-specific inflammatory processes.

Restricted junctional diversity of T cell receptor delta gene rearrangements expressed in systemic lupus erythematosus (SLE) patients.

SLE is an autoimmune connective tissue disorder affecting multiple organs, in which T cells may play a central role. This study investigated T cell receptor (TCR) gamma/delta repertoire expression in peripheral blood mononuclear cells (PBMC) of SLE patients and healthy individuals using variable (V) gene family-specific polymerase chain reaction (PCR) amplification of TCR cDNA. The expressed V gamma repertoires were diverse in SLE and control PBMC, although V gamma IV gene rearrangements were barely detectable or not expressed in some patients. In contrast, delta chain expression was limited in all SLE patients, with delta transcripts rearranged primarily to the V delta 1 and V delta 2 genes, as opposed to control PBMC, in which all six V delta genes were detected. To assess the clonality of TCR populations, cDNA clones containing rearranged V delta 1, V delta 2 and V gamma 9 transcripts were sequenced from PBMC of both patients and controls. For controls, delta chain junctional region sequences showed extensive molecular heterogeneity, since virtually all 34 V delta 1 and 32 V delta 2 cDNA clones analysed were unique. A few V gamma 9 cDNA clones (3/21) had the same junctional region sequence motif (EVQEL) encoded largely by the V gamma 9 and joining (J) gamma P gene segments. Identical V gamma 9 junctional sequences were found in SLE patients that did not contain the EVQEL motif present in normal peripheral blood gamma/delta lymphocytes. Moreover, the predominant V delta 1-J delta -constant (C) delta and V delta 2-J delta-C delta gene rearrangements expressed in SLE PBMC showed restricted junctional diversity, but the oligoclonal delta transcripts were different in each patient. These findings suggest in vivo oligoclonal expansion of gamma/delta T cells in the periphery of SLE patients in response to a limited number of nominal ligands. Whether gamma/delta T cells contribute to the development of systemic autoimmunity remains to be investigated.

Identification for human TCRgamma/delta‐recognized epitopes/proteins via CDR3delta peptide‐based immuno‐biochemical strategy

To identify antigenic epitopes/proteins recognized by TCR gamma/delta, we established a new immuno‐biochemical strategy, which is mainly based on binding activity of complementarity determining region 3 in TCR delta (CDR3delta). CDR3delta peptides were firstly synthesized. After validations for their binding specificity to target cells/tissues, the CDR3¦Ä peptides were employed as probes to pan putative epitopes in a 12‐mer random peptide phage‐displayed library, or to identify putative protein ligands within ovarian tumor protein extracts by CDR3¦Ä peptide‐mediated affinity chromatography and the analysis of LC ESI‐MS/MS. We identified nine peptides and two proteins for TCR gamma/delta, including human mutS homolog 2 (hMSH2) and heat shock protein 60. All nine tested epitope peptides could not only bind to gamma/deltaT cells, but also functionally activate gamma/deltaT cells in vitro. The hMSH2 on ovarian tumor cells was recognized by Vdelta2 T cells, suggesting that hMSH2 might be a new protein ligand for TCRdelta. Taken together, our findings provide a novel and facilitative immuno‐biochemical technical strategy to find epitopes/proteins recognized by gamma/deltaT cells.

Transcriptional activation of the cardiac homeobox gene CSX1/NKX2-5 in a B-cell chronic lymphoproliferative disorder

Homeobox containing transcription factors are frequently deregulated in human hematologic malignant diseases either indirectly through an abnormality of an upstream factor, or directly through rearrangement of the gene itself. Study of T-cell acute lymphoblastic leukemia identified the related non-clustered homeobox transcription factors, TLX1 and TLX3, as frequently ectopically expressed as a result of chromosomal translocations. We report the deregulation of a non-clustered homeobox gene in a new type of t(5;14)(q35;q11) translocation in a mature peripheral B-cell leukemia. This translocation results in the ectopic expression of the CSX1/NKX2-5 gene on chromosome 5q35 due to its juxtaposition to the TCR delta gene on chromosome 14q11. Expression of the CSX1/NKX2-5 protein conferred enhanced replating potential to transduced murine bone marrow cells. Our study establishes that deregulation of homeobox encoding genes is not restricted to acute leukemic proliferations, but is also observed in chronic malignant diseases.

T cells and cytokines in intracellular bacterial infections: experiences with Mycobacterium bovis BCG.

Intracellular bacteria reside in mononuclear phagocytes, and protective immunity is dominated by T lymphocytes. Mycobacterium bovis bacillus Calmette-Guéin (BCG) infection of mice represents an excellent model for studying immune mechanisms involved in defence against persistent intracellular bacteria that cause chronic disease. Gene disruption mutant mice include: A beta-/-, which lack conventional CD4+ T cell receptor alpha/beta (TCR alpha/beta) T lymphocytes; beta 2 microglobulin -/-, which lack conventional CD8+ TCR alpha/beta lymphocytes; TCR beta-/-, which lack all TCR alpha/beta lymphocytes; TCR delta-/-, which lack all TCR gamma/delta lymphocytes; and RAG-1-/- mutants, which lack mature T and B lymphocytes. Studies of these mutants suggest that CD4+ TCR alpha/beta, CD8+ TCR alpha/beta and TCR gamma/delta T lymphocytes all contribute to immunity against M. bovis BCG. Activation of antibacterial effector functions in macrophages by T helper 1 (Th1) cell-derived gamma-interferon (IFN-gamma) is central to protection. In contrast, Th2 cells are only marginally involved. Activation of Th1 and Th2 cells is regulated by interleukin 10 (IL-10) and IL-12, which are induced early in infection with M. bovis BCG. Although IL-12 is stimulated by M. bovis BCG in immunocompetent mice, studies with IFN-gamma receptor-deficient and tumour necrosis factor alpha (TNF-alpha) receptor-deficient mutant mice suggest that M. bovis BCG-induced IL-12 secretion depends on IFN-gamma and TNF-alpha. Hence, IL-12 cannot be the first cytokine produced during M. bovis BCG infection.

REGULATION OF THE POSTBURN WOUND INFLAMMATORY RESPONSE BY γδ T-CELLS

Healing of the burn injury site is a critical component of the patient's successful recovery from this form of trauma. Previous studies from our laboratory have demonstrated that gammadelta T-cells via the production of growth factors are important in burn wound healing. Nonetheless, the role of these cells in burn wound inflammation remains unknown. To study this, wild-type (WT) and gammadelta T-cell receptor-deficient (delta TCR) C57BL/6 male mice were subjected to burn injury or sham procedure. Wound cells were collected by implantation of polyvinyl alcohol sponges beneath the burn site in injured mice or beneath uninjured skin in sham mice. At 3 days after injury, infiltrating cells, wound fluid, and skin were collected for analysis. Burn injury markedly increased skin tumor necrosis factor-alpha (TNF-alpha) and monocyte chemoattractant protein 1 levels. In WT mice, the numbers of infiltrating cells were similar between nonburn wounds and burn wounds. In contrast, deltaTCRmice displayed a 6-fold reduction in the cellular infiltrate. Burn injury in WT mice caused a marked increase in burn wound TNF-alpha, monocyte chemoattractant protein 1, and interleukin 6 content as compared with nonburn wounds, whereas in delta TCRmice, the burn-induced increase of TNF-alpha and interleukin 6 was not observed. The wound cell infiltrate at 3 days postinjury was devoid of gammadelta T-cells in WT mice. It was predominately of myeloid origin expressing high levels of CD11b and F4/80. In conclusion, these findings suggest that resident gammadelta T-cells are important in the recruitment of inflammatory cells and regulation of the inflammatory response at the wound site after thermal injury.

Different chromosomal breakpoints impact the level of LMO2 expression in T-ALL

The t(11;14)(p13;q11) is presumed to arise from an erroneous T-cell receptor delta TCRD V(D)J recombination and to result in LMO2 activation. However, the mechanisms underlying this translocation and the resulting LMO2 activation are poorly defined. We performed combined in vivo, ex vivo, and in silico analyses on 9 new t(11;14)(p13;q11)-positive T-cell acute lymphoblastic leukemia (T-ALL) as well as normal thymocytes. Our data support the involvement of 2 distinct t(11;14)(p13;q11) V(D)J-related translocation mechanisms. We provide compelling evidence that removal of a negative regulatory element from the LMO2 locus, rather than juxtaposition to the TCRD enhancer, is the main determinant for LMO2 activation in the majority of t(11;14)(p13;q11) translocations. Furthermore, the position of the LMO2 breakpoints in T-ALL in the light of the occurrence of TCRD-LMO2 translocations in normal thymocytes points to a critical role for the exact breakpoint location in determining LMO2 activation levels and the consequent pressure for T-ALL development.

Highly restricted diversity of TCR delta chains of the amphibian Mexican axolotl (Ambystomamexicanum) in peripheral tissues

Gammadelta T cells localize at mammalian epithelial surfaces to exert both protective and regulatory roles in response to infections. We have previously characterized the Mexican axolotl (Ambystoma mexicanum) T cell receptor delta (TRD) chain. In this study, TRD repertoires in spleen, liver, intestine and skin from larvae, pre-adult and adult axolotls were examined and compared to the thymic TRD repertoire. A TRDV transcript without N/D diversity, TRDV1S1-TRDJ1, dominates the TRD repertoires until sexual maturation. In adult tissues, this canonical transcript is replaced by another dominant TRDV1S1-TRDJ1 transcript. In the thymus, these two transcripts are detected early in development. Our results suggest that gammadelta T cells that express the canonical TRDV1S1-TRDJ1 transcript emerge from the thymus and colonize the peripheral tissues, where they are selectively expanded by recurrent ligands. This particular situation is probably related to the neotenic state and the slow development of the axolotl. In thymectomized axolotls, TRD repertoires appear different from those of normal axolotls, suggesting that extrathymic gammadelta T cell differentiation could occur. Gene expression analysis showed the importance of the gut in T cell development.

Germ‐line and rearranged Tcrd transcription distinguish bona fide NK cells and NK‐like γδ T cells

NK cells and gammadelta T cells are distinct subsets of lymphocytes that contextually share multiple phenotypic and functional characteristics. However, the acquisition and the extent of these similarities remain poorly understood. Here, using T cell receptor delta locus-histone 2B-enhanced GFP (Tcrd-H2BEGFP) reporter mice, we show that germ-line transcription of Tcrd occurs in all maturing NK cells. We also describe a population of mouse NK-like cells that are indistinguishable from "bona fide" NK cells using standard protocols. Requirements for V(D)J recombination and a functional thymus, along with very low-level expression of surface TCRgammadelta but high intracellular CD3, define these cells as gammadelta T cells. "NK-like gammadelta T cells" are CD127+, have a memory-activated phenotype, express multiple NK cell receptors and readily produce interferon-gamma in response to IL-12/IL-18 stimulation. The close phenotypic resemblance between NK cells and NK-like gammadelta T cells is a source of experimental ambiguity in studies bridging NK and T cell biology, such as those on thymic NK cell development. Instead, it ascribes chronic TCRgammadelta engagement as a means of acquiring NK-like function.

Lymphome T gamma delta avec atteinte méningée exclusive révélé par une dermatomyosite

Introduction Peripheral T cell lymphomas are a heterogeneous group of post-thymic, mature lymphoid malignancies, accounting for approximately 10–15% of all non-Hodgkin's lymphomas. A rare entity within this group is represented by hepatosplenic T cell lymphoma, characterized by primary extranodal disease with infiltration of the liver and the spleen and by expression of the T cell receptor gamma delta chain. Exegesis A 64-year old man with dermatomyositis developed rapid-onset paraparesia and deafness. Cerebrospinal fluid analysis revealed large granular lymphomatous cells with CD3+ CD4– CD8– CD7+ CD16– CD56– surface antigens, expressing the gamma delta T-cell receptor. There was no evidence of skin or bone marrow infiltration by lymphoma or any other involvement. This is the first report of dermatomyositis associated with a gamma delta T-cell lymphoma (GDTL). Moreover, primitive and isolated meningeal involvement of such lymphomas has never been described before. Conclusion GDTL should be added to the differential list of neoplasia associated with dermatomyositis. Physiopathological mechanisms implicated in the neurological involvement of such lymphomas need to be elucidated.

Antigen recognition by γδ T cells

gammadelta T cells contribute to host immune competence uniquely. This is most likely because they have distinctive antigen-recognition properties. While the basic organization of gammadelta T-cell receptor (TCR) loci is similar to that of alphabeta TCR loci, there is a striking difference in how the diversity of gammadelta TCRs is generated. gammadelta and alphabeta T cells have different antigen-recognition requirements and almost certainly recognize a different set of antigens. While it is unclear what most gammadelta T cells recognize, the non-classical major histocompatibility complex class I molecules T10 and T22 were found to be the natural ligands for a sizable population (0.2-2%) of murine gammadelta T cells. The recognition of T10/T22 may be a way by which gammadelta T cells regulate cells of the immune system, and this system has been used to determine the antigen-recognition determinants of gammadelta T cells. T10/T22-specific gammadelta T cells have TCRs that are diverse in both V gene usage and CDR3 sequences. Their Vgamma usage reflects their tissue origin, and their antigen specificity is conferred by a motif in the TCR delta chain that is encoded by V and D segments and by P-nucleotide addition. Sequence variations around this motif modulate affinities between TCRs and T10/T22. That this CDR3 motif is important in antigen recognition is confirmed by the crystal structure of a gammadelta TCR bound to its ligand. The significance of these observations is discussed in the context of gammadelta T-cell biology.

T‐cell receptor gamma and delta gene rearrangements and junctional region characteristics in south Indian patients with T‐cell acute lymphoblastic leukemia

Clonal T-cell receptor (TCR) gamma and delta gene rearrangements were studied in 40 T-ALL cases (pediatrics, 29; adults, 11) using PCR with homo-heteroduplex analysis. At least one clonal TCRG or TCRD rearrangement was detected in 34 (85%) cases. TCR gamma (TCRG) rearrangement was detected in 25 (62.5%) cases that included 16 (55%) pediatrics and 9 (81.8%) adults. TCR delta (TCRD) rearrangement was detected in 14/40 (35%) cases, which included 12 (41%) pediatrics and 2 (18%) adults. The frequency of VgammaI-Jgamma1.3/2.3 was significantly more in adults than pediatrics (81.8% vs. 41.3%, P=0.02). In TCRD, Vdelta1-Jdelta1 was rearranged in 10 (25%) cases. The surface membrane CD3 positive cases are significantly associated with absence of TCRD rearrangements (surface membrane CD3+ TCRdelta- 84% vs. surface membrane CD3- TCRdelta- 48%, P value=0.03). Junctional region sequence analyzed with 10 cases each, of TCRG and TCRD, revealed an average junctional region of 7.4 nucleotides (range 2-18 nucleotides) in TCRG and 27 nucleotides (range 14-42 nucleotides) in TCRD-complete rearrangements. In TCRG, trimming at the ends of Vgamma and Jgamma germline nucleotides resulted in deletion, on an average of 9.2 nucleotides. In TCRD, deletion of nucleotides of the Vdelta and Jdelta gene segments on an average was 3.5 nucleotides. The junctional region of TCRD is more diverse than TCRG; nevertheless, the frequency of TCRG was more than that of TCRD and hence we rely more on TCRG clonal markers to quantitate the minimal residual disease in T-ALL.

Identification of a Subgroup of Biphenotypic Myeloid/T-Cell Acute Leukemias with a CEBPA-Mutant Gene Expression Profile, Frequent CEBPA Promoter Hypermethylation and NOTCH1 Mutations.

Recently, gene expression profiling has allowed the identification of specific molecular signatures of established as well as novel subgroups of acute myeloid leukemia (AML). CEBPA mutations are observed in ~10% of AML. In a gene expression profiling study of 285 AML cases, we previously found two distinct clusters consisting of almost all specimens with CEBPA mutations, i.e. cluster #15 and cluster #4. In cluster #4, only 9 out of 15 cases were CEBPA mutant. Thus, this cluster contained 6 patients with a highly similar expression signature, but without CEBPA mutations. We wished to investigate whether these cases represent a previously unidentified subgroup of leukemias. All 6 cases appeared morphologically immature (FAB-M0 or M1) and did not carry any of the common genetic aberrations. Supervised array expression analysis revealed CEBPA to be among the most highly downregulated genes in these 6 cases, which was associated with CEBPA CpG promoter hypermethylation in 4/6. None of the CEBPA mutant cases or a random selection of 10 AML cases from other clusters exhibited CEBPA promoter methylation. Strikingly, several T-cell related genes, e.g. CD7, CD3D, IL7R and T-cell receptor delta locus, appeared significantly overexpressed in these cases. Multicolor flowcytometry demonstrated high expression of CD34 and combinations of myeloid (e.g. CD13, CD33, CD117 and/or myeloperoxidase) as well as T-lymphoid (CD7, CD3, TdT) lineage markers on all six. Expression of T-cell markers correlated with T-cell receptor rearrangements in 4/6 cases. Other highly expressed genes in these biphenotypic acute leukemias included NOTCH1 and its recently identified target gene TRIB2. Sequencing analysis demonstrated putatively activating NOTCH1 mutations in three out of six samples, as described previously in T-cell acute lymphoblastic leukemia (T-ALL). The remaining 3 cases and 25 control AMLs, including all CEBPA mutant cases, did not demonstrate any NOTCH1 mutation. Finally, based on this first cohort of patients, we determined the minimal number of probe sets accurately predicting cluster #4 biphenotypic cases, and identified eight probe sets, including those for T-cell receptor delta/alpha locus. Applying this predictor set to an independent new cohort of 272 AML cases, an identical subgroup of another 6 AML patients with frequent NOTCH1 mutations and CEBPA promoter hypermethylation could be identified. Available clinical data suggested an adverse prognosis of this type of leukemia. In conclusion, we identified a subgroup of biphenotypic acute leukemias characterized by a CEBPA mutant gene expression profile, frequent CEBPA promoter hypermethylation, and frequent NOTCH1 mutations. To our knowledge, this is the first report to describe NOTCH1 mutations in leukemia other than T-ALL.

Hepatosplenic γδ T‐cell lymphoma following seven malaria infections

Hepatosplenic gammadelta T-cell lymphoma (HSTL) is a clinicopathological entity associated with an immunocompromised status in approximately 25% of patients. Herein is described a case of HSTL in a 53-year-old Brazilian man with seven previous malaria infections, initially misdiagnosed as a hyperreactive splenomegaly due to chronic malaria. A characteristic lymphoid infiltrate was observed in spleen, liver and bone marrow sinusoids/sinuses. Neoplastic cells had a CD45RO+, CD2+, CD7+, CD3+, CD5-, CD8+, CD56+, perforin+, FasL-negative, T-cell receptor (TCR)alphabeta-negative, TCRgammadelta+ profile. Analyses of gamma and delta TCR rearrangements confirmed diagnosis of gammadelta T-cell lymphoma by detecting VgammaI/Vdelta1-Jdelta1 clonal rearrangements. Sensitive polymerase chain reaction (PCR) for Plasmodium falciparum, Epstein-Barr virus and herpesvirus-8 failed to demonstrate infection. The disease progressed to a fatal outcome following cutaneous infiltration and leukemic proliferation. The authors also comment on the association of lymphoma and infection, focusing on PCR diagnosis of TCRgamma and delta clonal rearrangements and the presumed pathogenic events leading to HSTL in the context of chronic malaria infection. Initial lymphomagenic stages might not be direct consequences of antigenic stimulation of Vdelta1 T-cells, but might depend on interactions between gammadelta T and B cells during cooperative or regulatory responses to Plasmodium sp.

Hepatosplenic γδ T-cell lymphoma with ring chromosome 7, an isochromosome 7q equivalent clonal chromosomal aberration

Hepatosplenic T-cell lymphoma is a rare, clinically aggressive lymphoma. Most cases represent a neoplasm of mature non-activated gammadelta T cells. Isochromosome 7q i(7)(q10) is thought to be the primary cytogenetic abnormality of this disease. In this paper, we describe a hepatosplenic gammadelta T-cell lymphoma case, with clonal ring chromosome 7 exemplifying an isochromosome 7q equivalent clonal aberration. A 62-year-old female patient presented with thrombocytopenia, isolated hepatosplenomegaly, and extremely high levels of LDH. Bone marrow work-up demonstrated a sinusoidal cytotoxic T-cell infiltrate with blastic features, while molecular studies verified monoclonal rearrangement for both TCR gamma and TCR delta genes. Cytogenetics revealed clonal abnormalities including ring chromosome 7, trisomy 8, and der(19), while FISH analysis detected 7q amplification with partial deletion of 7p in ring chromosome 7. To the best of our knowledge, this is the first reported T-cell lymphoma case with ring chromosome 7.

Identification of three new bovine T-cell receptor delta variable gene subgroups expressed by peripheral blood T cells

To understand the biology of gammadelta T cells in ruminants, it is necessary to have a comprehensive picture of gammadelta T-cell receptor gene diversity and expression. In this study, three new subgroups of bovine T-cell receptor delta (TRD) variable genes were identified by RT-PCR and sequencing and homology with TRDV genes from other mammals determined. Previously unidentified TRDV subgroup genes described in this study include the bovine homologues of ovine TRDV2, TRDV3, and TRDV4 which were named accordingly. TRDV2 subgroup has two genes (TRDV2-1 and TRDV2-2) while we found the previously identified TRDV1 has at least eight genes corresponding to separate genomic sequences. Nucleotide and amino acid sequences for particular gene subgroups between cattle and sheep were more than 87% identical but identities among TRDV subgroups within a species were much less, with bovine TRDV4 having <45% identity to the other three bovine TRDV gene subgroups. Analysis of circulating bovine gammadelta T cells revealed that genes from all four TRDV subgroups were expressed in combination with TRDJ1, TRDJ3, and TRDC, although TRDV4 was the least represented, and all displayed a variety of CDR3 junctional lengths. Finally, some genes within the TRDV1, TRDV2, and TRDV3 subgroups recombined with TRAV incorporating TRAJs, suggesting dual use.

Characterization of the Immune Response in the Placenta of Cattle Experimentally Infected with Neospora caninum in Early Gestation

A serial examination of three groups of cattle infected intravenously (iv) (Group 1, n=8) or subcutaneously (sc) (Group 2, n=8) with live Neospora caninum tachyzoites or with VERO cells (Group 3, n=8) at 70 days' gestation was carried out and the nature of the inflammatory responses in the placenta and the presence of parasite antigen were analysed. Immune cells expressing CD3, CD4, CD8, gamma delta (gammadelta) T-cell receptors (TCR), CD79alpha cytoplasmic (cy) (B cells) and NKp46 [natural killer (NK) cells] antigens were identified immunohistochemically and cells expressing mRNA for interferon-gamma (IFN-gamma) were labelled by in-situ hybridization. Intravenous inoculation caused mortality in all fetuses from 28 days post-inoculation (dpi) onwards. Subcutaneous inoculation caused mortality in 50% of the animals by 28dpi. Pathological changes in the placenta consisted of necrosis of fetal placental villi, necrosis and inflammation in adjacent areas of the maternal septum and inflammation at the base of the maternal caruncle. The inflammatory infiltrate consisted mainly of CD3(+) lymphocytes, dominated by CD4(+) and gammadelta TCR(+) cells, with CD8(+) cells present to a lesser extent. The results from the control group indicated fewer NK cells than those occurring in the placenta of human beings or mice. Infiltration of CD4(+) cells and NKp46(+) cells was observed in the caruncular base and septa 14 days after infection, whereas infiltration of gammadelta TCR(+) cells was observed from 28 dpi onwards. To our knowledge this is the first report on the presence and distribution of NK cells in the bovine placenta. Maternal inflammatory cells expressing mRNA for IFN-gamma were identified in animals inoculated with parasites iv or sc at 14 and 28 dpi, respectively. In the sc-inoculated dams with live fetuses at 28, 42 and 56dpi, there was no evidence of parasite antigen, infiltration of immune cells or production of IFN-gamma, suggesting that the parasite had not reached the placenta. The exact cause of fetal death was not established. Tissue destruction by the parasite may have occurred; in addition, there may have been a T helper 1 (Th-1) immune response to the neospora infection at the materno-fetal interface, resulting in infiltrations of CD4T cells, gammadelta T cells and NK cells and the subsequent production of IFN-gamma. It is possible that a pro-inflammatory Th-1 response early in gestation protects the dam by eliminating the parasite; however, it may lead to destruction of the placental tissues themselves and thus be incompatible with fetal survival.