Genomic and phenotypic characterization of a red-pigmented strain of Massilia frigida isolated from an Antarctic microbial mat.

The McMurdo Dry Valleys of Antarctica experience a range of selective pressures, including extreme seasonal variation in temperature, water and nutrient availability, and UV radiation. Microbial mats in this ecosystem harbor dense concentrations of biomass in an otherwise desolate environment. Microbial inhabitants must mitigate these selective pressures via specialized enzymes, changes to the cellular envelope, and the production of secondary metabolites, such as pigments and osmoprotectants. Here, we describe the isolation and characterization of a Gram-negative, rod-shaped, motile, red-pigmented bacterium, strain DJPM01, from a microbial mat within the Don Juan Pond Basin of Wright Valley. Analysis of strain DJMP01's genome indicates it can be classified as a member of the Massilia frigida species. The genome contains several genes associated with cold and salt tolerance, including multiple RNA helicases, protein chaperones, and cation/proton antiporters. In addition, we identified 17 putative secondary metabolite gene clusters, including a number of nonribosomal peptides and ribosomally synthesized and post-translationally modified peptides (RiPPs), among others, and the biosynthesis pathway for the antimicrobial pigment prodigiosin. When cultivated on complex agar, multiple prodiginines, including the antibiotic prodigiosin, 2-methyl-3-propyl-prodiginine, 2-methyl-3-butyl-prodiginine, 2-methyl-3-heptyl-prodiginine, and cycloprodigiosin, were detected by LC-MS. Genome analyses of sequenced members of the Massilia genus indicates prodigiosin production is unique to Antarctic strains. UV-A radiation, an ecological stressor in the Antarctic, was found to significantly decrease the abundance of prodiginines produced by strain DJPM01. Genomic and phenotypic evidence indicates strain DJPM01 can respond to the ecological conditions of the DJP microbial mat, with prodiginines produced under a range of conditions, including extreme UV radiation.

Genome sequences of four agarolytic bacteria from the Bacteroidia and Gammaproteobacteria.

Here we present the genomes of four marine agarolytic bacteria belonging to the Bacteroidota and Proteobacteria. Two genomes are closed and two are in draft form, but all are at least 99% complete and offer new opportunities to study agar-degradation in marine bacteria.

Extracorporeal photopheresis as a curative treatment strategy in non epidermotropic T-cell lymphoma and large granular lymphocyte leukemia.

BACKGROUND: To evaluate the efficacy of extracorporeal photopheresis (ECP) in noncutaneous T-cell lymphoma and large granular lymphocytes leukemia (LGL). PATIENTS AND METHODS: We have treated 12 refractory/relapsed patients. Six peripheral T-cell lymphoma (PTCL), one T-lymphoblastic lymphoma and five LGL with blood involvement received six biweekly leukapheresis as induction phase, followed by one course a week for 4 weeks as consolidation and one course of maintenance per month for responders until progression/relapse or disappearance of the peripheral clone. RESULTS: Six patients responded to phototherapy. Two PTCL and two LGL achieved a complete response (CR) and two other PTCL a partial response. The median duration of CR was 117 months (45-150 months) for these four patients. The peripheral clone followed by flow cytometry decreased in all six responders. Two patients with a complete disappearance of the peripheral clone have not relapsed. CONCLUSIONS: As for cutaneous T-cell lymphoma, ECP therefore to be efficient for PTCL and LGL. Early decrease and disappearance of the peripheral clone were the indicators of clinical response and nonrelapse, respectively.

Efficient characterization of the TCR repertoire in lymph nodes by flow cytometry.

Analysis of the T-cell receptor (TCR) repertoire by flow cytometry proved to be relevant for investigating T-cell diversity and detecting reactive cells in blood samples. We used this approach to characterize non-malignant T-lymphocytes in lymph nodes and give insights into their origin. The TCR repertoire of CD4+ and CD8+ T-cells from 81 lymph nodes was analyzed with a four-color flow cytometer using a wide panel of 25 anti-Vbeta monoclonal antibodies. Flow cytometry proved to be a useful and informative technique. We demonstrated a diversified TCR-Vbeta repertoire, and only low level expansions, in 53% of the samples. They involved nearly all Vbeta families, were more frequent in the CD8+ subset of older patients, but were not related to pathology. No evidence could be demonstrated in favor of stimulation by common antigens. Interestingly, the TCR-Vbeta repertoire proved to be very similar in lymph nodes and blood samples. Our results argue that in the cases studied, lymph node enlargement is mainly due to an increased homing of circulating T-cells. They also provide reference values for expression of 25 TCR-Vbeta in lymph nodes, which could serve as a basis for further applications in diagnosis of T-cell lymphoproliferative disorders.

Immunophenotypic analysis of bone marrow B lymphocyte precursors (hématogones) by flow cytometry.

The aims of this flow cytometry study were to quantify B lymphoid precursors known as hématogones across age and clinical conditions and to study the immunophenotypic profile of these benign immature B cells. A total of 406 consecutive marrow specimens were analyzed for hématogones using 4-color flow cytometry during a 19 month period (60% males and 40% females). The age range was 3 months to 89 years. Hématogones were present in 80% of the specimens. Morphologic analysis of the smears from each patient showed small numbers of hématogones (<13% of total cellularity). The B cell population was defined by CD19 + CD45 bright positivity, coexpression of other B lineage markers: CD20, CD22, CD10, CD29, CD38 and CD58 in addition to HLA-DR and CD34. In our study we found a significant decline in hématogones with increasing age but a broad range was found at all ages. Marrow from some adults contained relatively high numbers. Diagnosis in these patients included cytopenias, infections, and neoplastic diseases. Distinction of hématogones is critical for disease management particularly after therapy of paediatric B acute lymphoblastic leukaemia to monitor for minimal residual disease.

Children with severe acute asthma admitted to Dutch PICUs: A changing landscape.

The number of children requiring pediatric intensive care unit (PICU) admission for severe acute asthma (SAA) around the world has increased. OBJECTIVES: We investigated whether this trend in SAA PICU admissions is present in the Netherlands. METHODS: A multicenter retrospective cohort study across all tertiary care PICUs in the Netherlands. Inclusion criteria were children (2-18 years) hospitalized for SAA between 2003 and 2013. Data included demographic data, asthma diagnosis, treatment, and mortality. RESULTS: In the 11-year study period 590 children (660 admissions) were admitted to a PICU with a threefold increase in the number of admissions per year over time. The severity of SAA seemed unchanged, based on the first blood gas, length of stay and mortality rate (0.6%). More children received highflow nasal cannula (P < 0.001) and fewer children needed invasive ventilation (P < 0.001). In 58% of the patients the maximal intravenous (IV) salbutamol infusion rate during PICU admission was 1 mcg/kg/min. However, the number of patients treated with IV salbutamol in the referring hospitals increased significantly over time (P = 0.005). The proportion of steroid-naïve patients increased from 35% to 54% (P = 0.004), with a significant increase in both age groups (2-4 years [P = 0.026] and 5-17 years [P = 0.036]). CONCLUSIONS: The number of children requiring PICU admission for SAA in the Netherlands has increased. We speculate that this threefold increase is explained by an increasing number of steroid-naïve children, in conjunction with a lowered threshold for PICU admission, possibly caused by earlier use of salbutamol IV in the referring hospitals.

Resistance to CD95-mediated apoptosis through constitutive c-FLIP expression in a non-Hodgkin's lymphoma B cell line.

CD95 (Fas/Apo-1) is a transmembrane molecule that induces apoptosis and plays a central role in the regulation of the immune response. The present study describes two new B lymphoid cell lines, B593 and BR97, derived from non-Hodgkin's lymphoma, which differ in susceptibility to CD95-mediated apoptosis. While B593 cells are sensitive to CD95mediated apoptosis, BR97 cells are completely resistant. Activation of caspase-8 and caspase-3 proteases plays an important role in the CD95 signalling pathway. CD95 stimulation induced caspase-8 and caspase-3 activation in B593, but not in BR97 cells. However, activation of both caspase-8 and caspase-3 was achieved in BR97 cells treated with staurosporine. Furthermore, protein synthesis inhibition by cycloheximide restored sensitivity to CD95-mediated apoptosis and allowed activation of both caspase-8 and caspase-3 in BR97 cells. These results indicate that, in BR97 cells, both caspases are functional and suggest that CD95-apoptosis resistance may result from the presence of inhibitory factor(s). Constitutive high level expression of the apoptotic inhibitor c-FLIP was observed in the CD95-resistant BR97 cell line compared to B593. Moreover, downregulation of c-FLIP expression level by protein synthesis inhibition strictly correlated with restored sensitivity to CD95-mediated apoptosis in BR97 cells. Furthermore, we demonstrate that c-FLIP is recruited to the CD95 DISC in BR97 cells together with caspase-8 and FADD. The data presented in this study strongly suggests that, in a B-NHL-derived cell line, resistance to CD95-mediated apoptosis results from endogenous high level expression of apoptotic inhibitor c-FLIP.

In situ leukemic plasmacytoid dendritic cells pattern of chemokine receptors expression and in vitro migratory response.

Plasmacytoid dendritic cell (PDC) leukemia/lymphoma is a rare neoplasm presenting cutaneous lesions at the time of diagnosis, followed by dissemination to bone marrow, lymph nodes, and other lymphoid and nonlymphoid organs. Since these leukemic counterparts of human PDC are similar to normal PDC, we studied their chemokine receptor equipment and their migratory capacities. We found both in skin lesions and in invaded lymph nodes an expression by tumor cells of CXCR3, CXCR4, and CCR7, and the concomitant expression by cells in the microenvironment of their respective ligands CXCL9, CXCL12, and CCL19. Moreover, flow cytometry phenotype of leukemic PDC (LPDC) revealed an unexpected expression of CCR6. We show that fresh tumor cells are able to migrate in response to CXCR4, CCR2, CCR5, CCR6, and CCR7 ligands, and the ability of CXCR3 ligands to increase the responsiveness to CXCL12. IL-3- or virus-induced activation of LPDC leads to downregulation of CXCR3 and CXCR4, and upregulation of CCR7, associated with the loss of response to CXCL12, and the acquisition of sensitivity to CCL19. Altogether, these results suggest that the preferential accumulation of LPDC in the skin or lymph nodes could be orchestrated by CXCR3, CXCR4, CCR6, and CCR7 ligands, found in nontumoral structures of invaded organs.

Differentiation of antigen-presenting cells (dendritic cells and macrophages) for therapeutic application in patients with lymphoma.

The recent clinical trial in lymphoma using tumor antigen-loaded DCs (Hsu et al, Nature Med 1996; 2: 52) demonstrates the efficiency of the use of professional antigen presenting cells (APCs) for taking up, processing and presenting tumor protein in a vaccine strategy in cancer. However, the production of large quantities of clinical grade APCs remains to be resolved. Here, we describe that both dendritic cells (DCs) and macrophages (MOs) can be efficiently differentiated in large numbers from lymphoma patients in spite of their disease and previous therapy. These cells were produced using the VAC and MAK cell processors according to standard operating procedures. DCs and MOs were differentiated from circulating monocytes in gas permeable hydrophobic bags, with 2% autologous serum and in the presence of GM-CSF and IL-13 or GM-CSF alone, respectively. DCs and MOs were then purified by counter flow centrifugation. Phenotypic, morphological and functional analysis showed that cells differentiated from patients with lymphoma present quite similar features to DCs and MOs produced from monocytes of healthy donors. Moreover, we show that MOs, when combined with CD20 antibody (Rituximab), can efficiently engulf tumor cells and propose that a such combination could be used for initiating a clinical trial in lymphoma. Thus, the possibility of producing functional DC and MOs in large amounts in conditions compatible with therapeutic application will allow the development of new immune strategies to eradicate lymphoma.

Quantification of cellular adhesion molecules on malignant B cells from non-Hodgkin's lymphoma.

The expression of five cellular adhesion molecules (CAMs), CD54, CD58, CD11a, CD29 and CD49d, was studied in 113 B cell non-Hodgkin's lymphomas (NHL) and in normal B cells from 12 control lymph nodes. Rather than reporting the percentage of positive cells, which does not discriminate between NHL subtypes, we quantified the intensity of CAM expression using flow cytometry. Apart from CD49d the expression of all these CAMs was statistically different among the NHL subtypes as defined by the REAL classification. Low grade NHL-small lymphocytic, follicular and mantle cell lymphoma--which are derived from quiescent cells and show an indolent disease course, expressed low levels of CAMs. Conversely, high grade NHL-diffuse large cell lymphoma--which are derived from proliferating cells and are clinically aggressive, expressed high levels of CAMs. These results indicate that in malignant NHL B cell tumour growth and clinical aggressiveness may be related to the adhesive capacities of the tumour cells.

Neutrophil activation by anti-beta 2 glycoprotein I monoclonal antibodies via Fc gamma receptor II.

Murine monoclonal antibodies (mAbs) to human beta 2-glycoprotein I (beta 2GPI), a plasma protein required for the binding of some antiphospholipid antibodies, have been shown to possess lupus anticoagulant properties and to activate platelets via Fc gamma receptor (Fc gamma R) crosslinking. Here we investigated their ability to induce polymorphonuclear leukocyte (PMN) functional responses. The six mAbs (IgG1 isotype) tested in combination with beta 2GPI led to a concentration-dependent activation of human PMNs as appreciated by granule release, H2O2 production, and cytosolic Ca2+ increase. This activation process was accompanied by the enhancement of PMN-mediated heparan sulfate loss from the endothelial cell line EA.hy 926 without evidence for cell lysis or detachment. F(ab')2 fragments of one of the mAbs bound to PMNs in a beta 2GPI-dependent manner but were devoid of activating effects. Carbamylated beta 2GPI was unable to mediate PMN-antibody binding and subsequent activation. In addition, cationization of beta 2GPI or removal of its sialic acid groups led to higher efficiency in binding to the PMN surface and triggering activation in comparison with the untreated protein. Thus, the process of PMN activation depends on mAb binding to these cells through both Fab (via beta 2GPI) and Fc domains, as confirmed by the suppression of all responses upon treatment with an anti-Fc gamma RII, but not anti-Fc gamma RIII, antibody. Our data suggest a model of cellular activation by beta 2GPI-dependent antiphospholipid antibodies.

Functional expression of CD80 and CD86 allows immunogenicity of malignant B cells from non-Hodgkin's lymphomas.

We analyzed the accessory function of malignant B cells from non-Hodgkin's lymphomas (NHLs). Among the 70 samples of malignant B cells included, four patterns of expression of the costimulatory molecules CD80 and CD86 were distinguished (+/+, +/-, -/+ and -/-). In two-thirds of the cases, CD80, CD86, or both were expressed. To investigate the relevance of these molecules for tumor immunogenicity, mixed lymphocyte reactions (MLR) were performed with allogeneic responding T cells and malignant B cells from nine NHL patients. Regardless of the level of expression of CD80 and CD86, significant proliferation was induced in the responder cells. The addition of monoclonal antibodies directed against CD80 and CD86 at the beginning of MLR almost completely inhibited this proliferation. We show that, during MLR, a high level of expression of CD80 and CD86 was induced in NHL B cells. Thus, cooperation between responding and stimulator cells seems to occur during MLR, allowing induction of optimal accessory function of B cells. We investigated whether malignant B cells cultured with CD40-L-transfected L cells in the presence of IL-4 could augment their antigen-presenting cell (APC) functions. The culture of NHL B cells in this sytem induced strong upregulation of the expression of CD80 and CD86 as well as other molecules involved in accessory cell functions (HLA class I, CD54, and CD58). In half of the cases, this activation resulted in enhanced proliferation of allo-T cells as compared to the proliferation induced by nonactivated malignant B cells. Our results show that NHL B cells are able to express functional CD80 and CD86 and to be fully competent APC. This suggests that the absence of an efficient T cell-mediated antitumor response in vivo is not related to a deficiency in the APC functions of malignant B cells.

Signal transduction via human leucocyte antigen class II molecules distinguishes between cord blood, normal, and malignant adult B lymphocytes.

Cord blood is increasingly used for hematopoietic stem cell transplantation since less severe graft-versus-host disease has been reported leading to the notion that cord blood is "naive." Human leucocyte antigen (HLA) class II molecules are expressed throughout B lymphocyte ontogeny (except the plasmocytes), are responsible for antigen presentation, and can also transmit signals. Cord blood B stimulate an allogeneic response, and this property is believed to indicate the presence of a class II-associated peptide. In this study we examined the capacity of cord blood B to transmit signals via HLA-DR. Activation and relocalization of protein kinase C (PKC) isoenzymes alpha and betaII was detected along with tyrosine kinase activation and proliferation. However, in contrast to resting adult B, generation of an intracellular calcium ([Ca++]i) flux and rapid aggregation were not detected. To address the question of whether or not HLA-DR signals throughout B lymphocyte ontogeny, we extended this study to include malignant adult B (B chronic lymphocytic leukemia [B-CLL], B mantle cell lymphoma, and B large cell leukemia). Tyrosine kinase activation and proliferation were observed in all these cell populations, albeit in the absence of [Ca++]i flux or an increase in PKC. HLA-DR therefore transmits signals throughout B lymphocyte ontogeny, although different signaling pathways are initiated in adult vs. fetal vs. malignant B. The lack of intracellular [Ca++]i flux in both cord blood and malignant B lymphocytes may represent a feature of HLA class II signaling at a particular stage of differentiation, although the downregulation of PKC clearly distinguishes between cord blood B and B-CLL.

Differentiation of antitumor-specific cytotoxic T lymphocytes from autologous tumor infiltrating lymphocytes in non-Hodgkin's lymphomas.

The present study describes a new culture protocol allowing the activation and proliferation of autologous tumor infiltrating T lymphocytes (TIL), and the generation of antitumor specific CTL in non-Hodgkin's lymphoma (NHL). Cells from eight patients with indolent NHL were used. We performed 3-week co-cultures of TIL with irradiated autologous malignant B cells in the presence of low doses of IL-1beta, IL-2 and IL-12. The proliferation, phenotype and cytotoxicity, and antitumor specificity of T cells recovered were studied. T-cell clonality was analyzed using TCRgamma gene rearrangement amplification by a multiplex PCR. Under these culture conditions, TIL proliferated, and the CD8+ T lymphocytes that were in a minority at the beginning of the culture increased dramatically in 6 out of 8 cases. In two cases, CD4+ T lymphocytes expanded. We showed that an oligoclonal selection of reactive T cells occurred in culture. Specific cytotoxicity developed against autologous malignant B cells in the 6 cases where there was an expansion of CD8+ T lymphocytes. Inhibition experiments performed with mAb directed against HLA class I and II molecules, CD4, CD8 and TCRgammadelta showed that the cytotoxic effector cells were CD8+ T lymphocytes probably expressing TCRalphabeta+. Cytokine secretion was analyzed in culture medium, and we detected significant levels of IFN-gamma, TNF-alpha, and IL-10 and no IL-4 (except in one case). Our results demonstrate that memory T cells from lymphoma patients can be amplified and differentiated into antitumor cytotoxic cells using a combination of the cytokines IL-1beta, IL-2, and IL-12 in association with non modified tumor cells.

Suppression of liver cell proliferation by glucocorticoid hormone: a comparison of normally growing and regenerating tissue in the immature rat.

The influence of glucocorticoid hormone on the time-course of liver regeneration in the immature rat has been studied by direct measurement of the rate of DNA accretion after the stimulus of partial hepatectomy. In contrast to hepatocyte proliferation associated with normal growth, which almost completely abolished by small doses of glucocorticoid, it is shown that even enormous amounts of hormone produce, at most, about a halving of the intrinsic cell proliferation rate in regenerating liver. Although deceptively magnified by the exponential growth pattern of the hepatic remnant, the inhibition of cell proliferation is thus considerably less complete than that induced in normally growing liver by much lower doses of hormone, a finding at distinct variance with the conclusions of earlier studies based entirely upon observations of radioactive precursor incorporation rather than direct measurement of DNA accretion. The mechanism by which a regenerative stimulus causes hepatocyte proliferation to lose its normal sensitivity to suppression by glucocorticoid, and thereby to exhibit a steroid insensitivity characteristic of other tissues in which cell proliferation reflects cell replenishment rather than normal growth, remains unknown.

Measurement of anti-phospholipid antibodies by ELISA using beta 2-glycoprotein I as an antigen.

The requirement of plasma cofactor beta 2-glycoprotein I for binding autoantibodies against anionic phospholipids has been reported. We describe the development of an enzyme-linked immunosorbent assay (ELISA) for anti-phospholipid antibodies using highly purified beta 2-glycoprotein I for coating microtitre plates. Intra- and interassay coefficients of variation, determined with serum pools of low, medium and high positivity, ranged between 3% and 18%. 54 sera from patients with systemic lupus erythematosus and related autoimmune disorders were analyzed by this assay; the results correlated well to those obtained in an ELISA using anionic phospholipids on the solid phase (r = 0.85, P less than 0.001). The two ELISA systems showed similar sensitivities although 8/31 positive sera scored negative in the beta 2-glycoprotein I ELISA. The latter group of eight sera showed significantly higher anti-phosphatidylcholine/anti-phosphatidylserine binding ratios than the group of 23 sera which scored positive in both assays. This new assay should permit accurate measurement of most of the clinically relevant anti-phospholipid antibodies and avoid inconsistencies likely to arise from secondary interactions that characterize lipid-based ELISA.

Staphylococcal protein-A is a potent T-cell mitogen of tonsillar lymphocytes.

Optimal conditions for the stimulation of human tonsillar lymphocytes by staphylococcal protein-A (SpA) are described. By stimulating fractions enriched or depleted of E-rosette forming cells, the response was shown to be predominantly a T-cell response. A comparison of stimulation by SpA with that of phytohaemagglutinin (PHA) proved SpA to be a potent T-cell mitogen. We suggest that SpA may be another useful mitogen for studying human T-cell growth and differentiation.

Impaired clonogenic potential of CD25 positive T cells in lymph nodes involved by B cell non-Hodgkin's lymphomas.

In vivo activated T cells (CD25+) present in lymph nodes involved by B-non-Hodgkin's lymphomas (B-NHL) were investigated here for their ability to proliferate in vitro. CD25-/CD25+ T cells were isolated using a rosette method with magnetic beads, then the frequency of proliferating T lymphocyte-precursors (PTL-P) in both populations was assessed by limiting dilution experiments, in the presence of IL2, PHA and allogeneic spleen cells as feeders. In a total of 16 cases studied, growing microcultures were observed in all cases for CD25- T cells (mean value of PTL-P frequency: 1/32; range 1/10 - 1/2899) but in 6 cases only for CD25+ T cells (mean value of PTL-P frequency: 1/441; range 1/119 - 1/3736); the absence of any proliferative cultures in the 10 other cases indicated that the number of PTL-P was inferior to 1/12480. These results suggest that the proliferative potential of CD25+ T cells infiltrating lymph nodes involved by B-NHL is paradoxically decreased.

Limiting dilution analysis of the frequency of IL2 responsive T cells in lymph nodes involved by B-cell non-Hodgkin's lymphomas.

Total T lymphocytes separated from twelve lymph nodes involved by B-NHL were studied in limiting dilution experiments for their ability to proliferate in the presence of both R-IL2 used at a final concentration of 40 U/ml and irradiated autologous malignant B cells as feeders. The number of proliferating T-lymphocyte precursors (PTL-P) thus estimated was low in each case (mean: 1/4503; range, 1/200 to 1/11013). Once expanded, proliferation of the IL2 responsive T cells in the presence of autologous malignant B cells remained strictly dependent on the addition of exogenous IL2. Control cases consisted of T lymphocytes separated from peripheral blood of six healthy subjects and cultured in the presence of both R-IL2 (40 U/ml) and irradiated autologous total mononuclear cells as feeders; the mean frequency of PTL-P thus obtained (1/173; range, 1/49 to 1/457) was significantly higher than in malignant lymph nodes (p less than 0.01). These findings do not support the hypothesis that, in this series of patients, expansion of malignant B cells may lead to the activation and growth of T cells sensitized against the tumour.

Immunotherapy by non-myeloablative stem cell transplantation: study of the immune reconstitution. Arguments for distinct cell subsets in skin and blood.

INTRODUCTION: Non-myeloablative peripheral stem cell transplantation has been shown to induce tumour rejection in patients with acute leukaemia. However, the immunological mechanisms involved and the immune reconstitution achieved have not been investigated. MATERIALS AND METHODS: We describe the cases of two patients for whom we have studied the lymphocyte reconstitution achieved, using both phenotypic and genetic analyses of the T-cell repertoire, after peripheral stem cell transplantation. RESULTS: : In both cases we observed immune reconstitution with T-cell repertoire evolution and presence of activated CD8(+) T cells. In one of the patients an activated clone expressing Vbeta8 represents 46% of the CD8(+) cells. Expansion of this clone occurred in the absence of graft vs host disease symptoms. In the second case a skin lesion typical of graft vs host disease appeared after complete remission had been achieved. The T-cell repertoire in a biopsy of the lesion was distinct from that observed in the blood. CONCLUSION: Our study indicates that peripheral donor cells can effectively reconstitute a grafted patient while inducing an immune response against antigens expressed by the leukaemic/myeloma cells. Our data provide arguments for different populations of T cells associated with graft vs leukaemia/lymphoma and GVH effects.