Activation of iNKT cells by a distinct constituent of the endogenous glucosylceramide fraction

Invariant natural killer T (iNKT) cells are a specialized T-cell subset that recognizes lipids as antigens, contributing to immune responses in diverse disease processes. Experimental data suggests that iNKT cells can recognize both microbial and endogenous lipid antigens. Several candidate endogenous lipid antigens have been proposed, although the contextual role of specific antigens during immune responses remains largely unknown. We have previously reported that mammalian glucosylceramides (GlcCers) activate iNKT cells. GlcCers are found in most mammalian tissues, and exist in variable molecular forms that differ mainly in N-acyl fatty acid chain use. In this report, we purified, characterized, and tested the GlcCer fractions from multiple animal species. Although activity was broadly identified in these GlcCer fractions from mammalian sources, we also found activity properties that could not be reconciled by differences in fatty acid chain use. Enzymatic digestion of β-GlcCer and a chromatographic separation method demonstrated that the activity in the GlcCer fraction was limited to a rare component of this fraction, and was not contained within the bulk of β-GlcCer molecular species. Our data suggest that a minor lipid species that copurifies with β-GlcCer in mammals functions as a lipid self antigen for iNKT cells.The lines that distinguish innate from adaptive immunity are less clear than once believed. It is now appreciated that innate T cells, including invariant natural killer T (iNKT) cells, mucosal associated invariant T (MAIT) cells, γδ T cells, and some CD1a/b/c restricted T cells, constitute a substantial portion, perhaps 10–20% of the normal human T-cell repertoire (1). Innate T cells use the machinery of the adaptive immune system to generate antigen receptors with limited diversity and conserved antigenic specificity. Identification of the antigenic targets recognized by these cells is a fundamental step in understanding their biology in health and disease.As the best studied member of the innate T-cell family, iNKT cells have been shown to play a role in a variety of immune contexts, including inflammatory processes such as autoimmunity and allergic disease, cancer surveillance, and prominently, in host defense to bacteria, fungi, and viruses (2). In most situations, the activation of iNKT cells requires the recognition of stimulatory lipid antigens presented by a specialized antigen-presenting molecule named CD1d. The first lipid antigen discovered, α-galactosylceramide, was purified from a marine sponge, and the α-galactose headgroup was determined to be critical for antigenicity (3). Mammals have not been shown to produce gluco- or galactolipids where the headgroup is attached with α stereochemistry. Following the discovery of α-galactosylceramide, several microbes, including Sphingomonas species (4–6), Borrelia burgdorferi (7), and Streptococcus pneumoniae (8), were shown to produce α-anomeric lipid antigens, suggesting that α-glycolipids could be a microbial signature.Although microbe-derived lipids are likely to contribute to iNKT cell activation during bacterial and fungal infections, iNKT cells also play an important role in multiple diseases where foreign lipids are not expected to be present, such as viral infection, autoimmunity, and cancer, or in response to Toll-like receptor agonists (9). These observations suggested that endogenous lipid antigens play a central role in activating iNKT cells during many immune responses. Several candidate endogenous lipid antigens have been identified, including isoglobotrihexosylceramide (iGb3) (10), lyso-phosphatidylcholine (11), and plasmalogen lyso-phosphatidylethanolamine (12). We have reported activity in mammalian glucosylceramides (GlcCers), and we implicated β-glucosylceramides that are widely found in mammalian tissues (13). Of note, none of these endogenous antigens were described to contain an α-anomeric glycolipid.In this report, we characterized the structures and activities present in GlcCer-enriched lipid fractions from multiple sources, including those from the milk and serum of multiple animal species. Although the GlcCer fractions from multiple origins activated iNKT cells, we identified a GlcCer-enriched lipid fraction from a human source that was unable to activate iNKT cells despite containing GlcCers with similar molecular composition to activating GlcCers from other sources. This led us to consider the possibility that activity in the naturally occurring mammalian GlcCer fractions was contributed by a rare component of the total GlcCer species. We used multiple enrichment strategies to purify the activity in the GlcCer fraction to a rare component of the starting material. The functional, mass spectrometry (MS), and NMR spectroscopy characterization of this activity is reported herein.     

CD4-positive T-helper cell responses to the PASD1 protein in patients with diffuse large B-cell lymphoma

Background

Vaccine development targeting the novel immunogenic Per ARNT Sim Domain containing 1 (PASD1) cancer testis antigen represents an attractive therapeutic approach for the significant number of patients with diffuse large B-cell lymphoma who are refractory to conventional treatment. Since CD4-positive T helper cells have crucial roles in promoting and maintaining immune responses to tumor antigens, the presence of a CD4-positive T-helper immune response to the PASD1 antigen in patients with diffuse large B-cell lymphoma was investigated in the current study.

Design and Methods

Thirty-one patients with diffuse large B-cell lymphoma (25 with de novo, five with transformed and one with T-cell-rich B-cell lymphoma) were studied. Five immunogenic PASD1 peptides predicted to bind to several major histocompatibiliy complex, class II DR beta 1 alleles were identified using web-based algorithms. Peripheral blood mononuclear cells from patients were used to investigate the immunogenicity of these DR beta 1-restricted peptides in vitro using both gamma-interferon release enzyme-linked immunospot and cytolytic assays.

Results

Two of the five PASD1 peptides, PASD1(6) and PASD1(7), were shown to be immunogenic in 14 out of 32 patients studied in a gamma-interferon release assay. CD4-positive T-helper cell lines from two patients raised against PASD1 peptides were able to lyse cell lines derived from hematologic malignancies expressing endogenous PASD1 protein.

Conclusions

This is the first report of a CD4-positive T-helper response to the PASD1 protein in patients with lymphoma. The immunogenic peptides described here represent valuable additional candidates for inclusion in a vaccine to treat patients with PASD1-positive diffuse large B-cell lymphoma whose disease is refractory to conventional therapies.     

Comparable T helper 1 (Th1) and CD8 T-cell immunity by targeting HIV gag p24 to CD8 dendritic cells within antibodies to Langerin, DEC205, and Clec9A

Improved protein-based vaccines should facilitate the goal of effective vaccines against HIV and other pathogens. With respect to T cells, the efficiency of immunization, or "immunogenicity," is improved by targeting vaccine proteins to maturing dendritic cells (DCs) within mAbs to DC receptors. Here, we compared the capacity of Langerin/CD207, DEC205/CD205, and Clec9A receptors, each expressed on the CD8(+) DC subset in mice, to bring about immunization of microbial-specific T cells from the polyclonal repertoire, using HIV gag-p24 protein as an antigen. α-Langerin mAb targeted splenic CD8(+) DCs selectively in vivo, whereas α-DEC205 and α-Clec9A mAbs targeted additional cell types. When the mAb heavy chains were engineered to express gag-p24, the α-Langerin, α-DEC205, and α-Clec9A fusion mAbs given along with a maturation stimulus induced comparable levels of gag-specific T helper 1 (Th1) and CD8(+) T cells in BALB/c × C57BL/6 F1 mice. These immune T cells were more numerous than targeting the CD8(-) DC subset with α-DCIR2-gag-p24. In an in vivo assay in which gag-primed T cells were used to report the early stages of T-cell responses, α-Langerin, α-DEC205, and α-Clec9A also mediated cross-presentation to primed CD8(+) T cells if, in parallel to antigen uptake, the DCs were stimulated with α-CD40. α-Langerin, α-DEC205, and α-Clec9A targeting greatly enhanced T-cell immunization relative to nonbinding control mAb or nontargeted HIV gag-p24 protein. Therefore, when the appropriate subset of DCs is targeted with a vaccine protein, several different receptors expressed by that subset are able to initiate combined Th1 and CD8(+) immunity.     

Activation of autologous leukemia-specific T cells in acute myeloid leukemia: monocyte-derived dendritic cells cocultured with leukemic blasts compared with leukemia-derived dendritic cells

In acute myeloid leukemia (AML) blasts can be differentiated into dendritic cell (DC) like cells (AML-DC). These cells have a mature DC-like phenotype, are strong stimulators in mixed leukocyte reactions and can be used to generate leukemia-specific cytotoxic T cells. However, recent reports about naturally existing leukemic DC with immunoregulatory dysfunctions in peripheral blood of AML patients caused concerns about the use of AML-DC for therapeutic purposes. Systematic intra-individual comparisons between AML-DC and non-leukemic DC derived from monocytes (MoDC) in AML patients are missing. Thus, we investigated the ability to generate MoDC from peripheral blood of 17 AML patients in first remission and their functional integrity to stimulate leukemia-specific T cells by simple coculture with leukemic blasts. Phenotypic analysis of AML-DC and MoDC from the same individual patients revealed that MoDC exhibit a more homogenous mature DC phenotype. Additionally, functional analysis demonstrated the ability of remission MoDC to activate autologous leukemia-specific T cells in 11 of 12 patients, whereas AML-DC led to a specific T cell activation in four of eight patients. The presented findings might have impact on the design of further therapeutic studies using autologous antigen-presenting cells.     

Clonal CD8+ and CD52– T cells are induced in responding B cell lymphoma patients treated with Campath-1H* (anti-CD52)

Abstract: Five patients with non-Hodgkin's lymphoma (NHL) and 4 patients with chronic lymphocytic leukaemia (CLL) were treated with the CDR-grafted (rat × human) monoclonal antibody (mAb) Campath-1H (anti-CD52). Tumour regression was noted preferentially in peripheral blood and in the bone marrow but lymph nodes were less affected. Normal blood B and T cells were profoundly reduced in all patients whereas CD16⁺ NK cells and CD14⁺ monocytes decreased marginally. In all responding CLL patients CD52-negative T but not B cells appeared during treatment and persisted for several months (4–19+) during unmaintained follow-up. Clonal T cells defined as a predominance of a single T cell receptor (TCR) V gene usage, in one case verified by TCR CDR3 fragment analysis and nucleotide sequencing, emerged within the CD52^–/CD8⁺ cell population during Campath-1H therapy in 2 CLL patients, both achieving a long-lasting remission. The increase in CD8⁺ T cell expansions (up to 23-fold) during unmaintained remission and follow-up suggest that the clonal CD8⁺ cells may represent regulatory T cells controlling the growth of the tumour B cell clone. Clonal T cells might thus be a target for an immune therapeutic intervention in B cell tumours.     

Increase in CD95 (Fas/APO-1)-positive CD4+ and CD8+ T cells in peripheral blood derived from patients with autoimmune hepatitis or chronic hepatitis C with autoimmune phenomena

BACKGROUND: The expressions of CD95 (Fas/APO-1) and Bcl-2 are determinants of apoptosis in normal lymphocytes, and abnormalities in their expressions might contribute to the induction of autoimmunity. In this study, we examined the expressions of CD95 and Bcl-2 on freshly isolated T and B cells from patients with autoimmune hepatitis (AIH) or chronic hepatitis C associated with autoimmune phenomena (CH-C(AI)). METHODS: The CD95 and Bcl-2 expressions within CD4+ T, CD8+ T, and CD19+ B cell subsets were analysed by two-colour flow cytometry. RESULTS: The surface expression of CD95 was significantly high in both the CD4+ T and CD8+ T cell subsets derived from the patients with AIH and those with CH-C(AI), compared with expression in patients with CH-C and normal subjects. The increase in CD95 expression was associated with the phenotypic conversion of naive CD45RO- to primed CD45RO+ CD4+ T cells. Bcl-2 was detected in the vast majority of peripheral T and B cells. There was no significant difference in the percentage of Bcl-2-positive cells in the CD4+ T cell, CD8+ T cell and CD19+ B cell subsets among the patient groups and normal subjects. CONCLUSIONS: These results indicate that an increase in CD4+ T cells expressing CD45RO and CD95 marks an important subset of AIH and CH-C(AI) patients. These expanded CD95+ CD45RO+ primed T cells most likely reflect a continuous antigen-specific or non-specific activation of T lymphocytes, and/or the persistent presence of activated lymphocytes as a consequence of abnormalities in the peripheral deletion of activated lymphocytes. These persistently activated lymphocytes might play a role in the induction of autoimmunity in AIH and CH-C(AI).     

Loss of Ia-positive epidermal Langerhans cells at the onset of Type 1 (insulin-dependent) diabetes mellitus

Summary Immunocompetent antigen-presenting Langerhans cells were investigated in skin biopsies of 20 short-term Type 1 (insulin-dependent) diabetic patients and compared with 17 matched normal control subjects. Langerhans cells in epidermal sheet preparations were visualized with a monoclonal anti-HLA DR antibody using indirect immunofluorescence. A significant decrease of Langerhans cells/mm² body surface area was found in 10 patients immediately at the onset of diabetes compared to 10 patients with 6 months duration of diabetes and to normal control subjects (401±30 vs 559±43 vs 611±33, p<0.01 and p<0.002). There was no significant difference in the number of Langerhans cells between patients with 6 months duration of diabetes and control subjects. Examination of the most likely precursor of Langerhans cells, the blood monocytes, indicated an increase of monocyte counts in Type 1 diabetic patients after 6 months duration (344±37 cells/l vs 191±31 in control subjects, p<0.05) and an inverse correlation between the number of Langerhans cells in skin with the number of monocytes in peripheral blood (at onset: r=–0.73, p<0.01, after 6 months of diabetes: r=–0.61, p<0.05). In addition, a positive correlation between Langerhans cells and daily insulin dose was noted in patients after 6 months of diabetes (r=0.76, p<0.01). The data suggest a loss of Langerhans cells in skin at the onset of Type 1 diabetes and that functional alterations of these and perhaps also other antigenpresenting cells may be involved in the pathogenesis of Type 1 diabetes.     

Treatment of acute lymphoblastic leukaemia with the second generation of CD19 CAR‐T containing either CD28 or 4‐1BB

T cells modified with anti‐CD19 chimeric antigen receptor (CAR) containing either CD28 or 4‐1BB (also termed TNFRSF9, CD137) costimulatory signalling have shown great potential in the treatment of acute lymphoblastic leukaemia (ALL). However, the difference between CD28 and 4‐1BB costimulatory signalling in CAR‐T treatment has not been well elucidated in clinical trials. In this study, we treated 10 relapsed or refractory ALL patients with the second generation CD19 CAR‐T. The first 5 patients were treated with CD28‐CAR and the other 5 patients were treated with 4‐1BB CAR‐T. All the 10 patients were response‐evaluable. Three patients achieved complete remission and 1 patient with extramedullary disease achieved partial response after CD28‐CAR‐T treatment. In the 4‐1BB CAR‐T treatment group, 3 patients achieved complete remission. Furthermore, FLT‐3 ligand (FLT3LG) was highly correlated with response time and may serve as a prognosis factor. No severe adverse events were observed in these 10 treated patients. Our study showed that both CD28 CAR‐T and 4‐1BB CAR‐T both worked for response but they differed in response pattern (peak reaction time, reaction lasting time and reaction degree), adverse events, cytokine secretion and immune‐suppressive factor level.     

Proliferatory defect of invariant population and accumulation of non-invariant CD1d-restricted natural killer T cells in the joints of RA patients

Abstract

Objectives. While numerical and functional defects of invariant NKT cells have been demonstrated in rheumatoid arthritis (RA), the detailed characterization of proliferative and secretory responses following CD1d-mediated presentation is lacking; the presence of non-invariant populations has never been assessed in human autoimmunity. We have evaluated both invariant and non-invariant populations in the blood and synovial fluid from patients to assess feasibility of NKT cell-directed manipulations in RA.

Methods. NKT cell populations were quantified by anti-CD4/anti-Vα24 staining and/or CD1d tetramers. Proliferation was measured in cultures of mononuclear cells following stimulations with αGalCer and cytokine secretion determined by multi-bead assay.

Results. We have confirmed a proliferative defect of iNKT cells in both peripheral blood and synovial fluid from RA patients, but no changes in baseline frequencies. Moreover, we have detected an enlargement of non-invariant cell pool in synovial fluid samples. In addition, we noted an evident Th2 shift following exposure to αGalCer and pronounced IL-6 secretion.

Conclusions. While RA patients suffer from defective proliferative responses of invariant NKT cells, non-invariant cells accumulate at the site of inflammation. While stimulation with αGalCer results in reduced TNF-α and increased suppressive IL-10, abundantly produced IL-6 could potentially contribute to the induction of Th17 cells in the joints.     

Primary CD30 (Ki-1)-positive anaplastic large cell lymphoma of the duodenum

Summary Ki-1 anaplastic large cell lymphoma (ALCL) commonly affects the skin, lymph nodes, and bone. Primary ALCL of the alimentary tract is rare. The authors describe a case of primary ALCL of the duodenum in a 62-year-old man who presented with epigastric discomfort and weight loss. The patient was treated with modified CHOP and is in complete remission at 18 months. We reviewed all cases of primary ALCL of the alimentary tract in the literature and noted that this neoplasm often mimics gastrointestinal carcinoma.     

Fas/APO-1 (CD95)-mediated cytotoxicity is responsible for the apoptotic cell death of leukaemic cells induced by interleukin-2-activated T cells

Apoptotic cell death is induced by the cross-linking of Fas/APO-1 receptor (CD95) in acute myelogenous leukaemia (AML) cells. Since CD95 ligand (CD95L) is expressed on interleukin-2 (IL-2)-activated T cells, we investigated the involvement of CD95-CD95L pathway in T cell-mediated cytotoxicity against AML cells. Activated CD8⁺ T cells could efficiently kill a parental CD95-sensitive AML cell line, MML-1 and, to a lesser extent, a CD95-resistant clone, MML-1R. Neither MML-1 nor MML-1R cells were killed by activated CD4⁺ T cells. The blocking monoclonal antibody (MoAb) against CD95, ZB4, caused a significant inhibition of T-cell-mediated cytotoxicity against MML-1 cells, but not against MML-1R cells. Interestingly, MML-1 cells underwent the classic nuclear morphologic changes and DNA fragmentation characteristic of apoptosis when cultured with activated T cells. Enumeration of apoptotic and necrotic nuclei showed that both apoptosis and necrosis were induced in MML-1 cells, whereas necrosis was exclusively observed in MML-1R cells. Apoptosis of MML-1 cells was completely blocked in the presence of ZB4 MoAb. Similarly, blocking by ZB4 MoAb significantly inhibited T-cell-mediated lysis of fresh AML cells in a CD95-sensitive group, but not in a CD95-refractory group. In addition CD8⁺ T cells expressed CD95L mRNA more abundantly than CD4⁺ T cells upon activation by IL-2. These findings suggest that T-cell-mediated cytotoxicity against AML cells requires participation of CD95-CD95L pathway for cytotoxic signal transduction leading to target apoptosis.     

Circulating tumor cells counts are associated with CD8+ T cell levels in programmed death-ligand 1-negative non-small cell lung cancer patients after radiotherapy: A retrospective study

This study aimed to explore the dynamics of circulating tumor cells (CTCs) and CD8+ T cells in stage II–III non-small cell lung cancer patients with CTCs in different programmed death-ligand 1 (PD-L1) status treated with radiotherapy and evaluate the correlation between CTCs and CD8+ T cells.This study was a retrospective study which reviewed 69 stage II–III non-small cell lung cancer patients underwent postoperative radiotherapy and peripheral blood tests of CTCs and T lymphocyte were available before radiation, 1 week after radiation and 1 month after radiation.In this study, 25 patients had PD-L1 positive CTCs and 44 patients had PD-L1 negative CTCs. The CTCs count was significantly decreased compared with baseline in patients with different PD-L1 status CTCs at 1 week and 1 month after radiotherapy. The proportion of CD8+ T cells was significantly increased at 1 month after radiotherapy compared with baseline in the total population (mean change, 7.24 ± 2.12; P < .05) and patients with PD-L1 negative CTCs (mean change, 7.17 ± 2.65; P < .05). One month after radiotherapy, the proportion of CD8+ T cells was negatively correlated with the CTCs count in the total population (r = −0.255, P = .034) and PD-L1 negative patients (r = –0.330, P = .029). In patients with PD-L1 negative CTCs, the CTCs count 1 week after radiotherapy (hazard ratio, 0.150 [95% confidence intervals., 0.027–0.840], P = .031) and the proportion of CD8+ T cells 1 month after radiotherapy (hazard ratio, 7.961 [95% confidence intervals, 1.028–61.68], P = .047) were independent prognostic factors for disease recurrence.After radiotherapy, only PD-L1-negative patients had a significant increase in the CD8+ T cell levels, while it was negatively correlated with CTCs count and was an independent prognostic factors of disease recurrence.     

Anaplastic large cell lymphoma (CD30+/Ki-1+): results of a prospective clinico-pathological study of 69 cases

Summary. Sixty-nine anaplastic large cell lymphomas (ALCLs) were selected from an Italian comparative trial on MACOP-B and F-MACHOP. As no significant difference in effectiveness of the protocols emerged, they were considered homogenously treated. The ALCLs were divided into two groups according to previously defined criteria: 41 were common type (ALCLs-CT) and 28 Hodgkin-related (ALCLs-HR). T-cell phenotype was most common (58%), while B-cell, null and hybrid forms accounted for 27%, 13% and 2%. Clinically, ALCLs CT and HR differed as to mean age (27 v 34·3 years) and presentation; all ALCLs-HR showed mediastinal involvement, with bulky disease in 57%, and more frequent occurrence in stage II. In contrast, ALCLs-CT showed mediastinal masses in 58·5%, infrequently revealed bulky disease (24%), and were not specifically associated to stage. Among the ALCLs-CT, 68·4% achieved complete remission (CR), 24·4% partial remission (PR), one (2·4%) was resistant to therapy, and two (4·8%) had fatal drug toxicity. Of the ALCLs-HR, 67·8% reached CR, 14·3% PR, and 17·9% did not respond. In CR, ALCLs-CT showed a greater tendency to relapse (32·1% v 14·2%). At present, 65·8% of ALCLs-CT and 67·8% of ALCLs-HR are alive with overall survival/disease-free survival averages of 31/27 and 29/24 months respectively. Our data emphasize that, independently of subtype, ALCLs benefit from the application of third-generation protocols for high-grade non-Hodgkin's lymphomas.     

Generation of HLA-DRB1*1501-restricted p190 minor bcr-abl (e1a2)-specific CD4+ T lymphocytes

A small population of cells in acute lymphoblastic leukaemia is characterized by a specific translocation of the c-abl oncogene on chromosome 9 to the break point cluster lesion (bcr) on chromosome 22, t(9; 22)(q34; q11) (e1a2). Theoretically, the junction-spanning sequences of oncogene fusion proteins might be ideal targets for immunotherapy because these are not present in normal cells. In this study, we show for the first time that in vitro immunization with a 17-mer e1a2 peptide representing the p190 minor bcr-abl fusion protein resulted in HLA-DRB1*1501-restricted peptide-specific proliferative CD4+ T lymphocytes, using peptide-pulsed monocyte-derived dendritic cells as the antigen-presenting cells.     

Co-expression of CD30 ligand and interleukin 4 (IL-4) receptors by acute myeloid leukaemia blasts is associated with the expansion of IL-4-producing CD30+ normal T cells

CD30 ligand (CD30L), but not its cognate receptor CD30, is frequently expressed on acute myeloid leukaemia (AML) blasts. In the present study, we found that leukaemic blasts presenting surface CD30L displayed a characteristic cytokine-receptor pattern that makes them ideal targets for those cytokines usually produced by Th2-type cell subsets. In particular, even though a broad distribution of Th2 cytokine receptors by AML blasts was shown, we demonstrated the almost exclusive expression of interleukin 4 (IL-4) receptor (R), in the absence of its cognate cytokine, by CD30L+ AML. Furthermore, a number of Th2-associated markers, including CD30, IL-4 and GATA-3, were expressed by residual T cells derived from CD30L+ AML but not from CD30L- AML, in which the presence of the Th1-associated marker LAG-3 was documented in some cases. The production of IL-4 in the absence of interferon gamma (IFN-gamma) was also detected in CD3+/CD30+ T cells from CD30L+ AML. These results, along with the shift toward IL-4-producing specific T-cell clones observed in CD30L+ AML samples by enzyme-linked Immunospot (ELISpot) assay, were consistent with the hypothesis of a Th2 polarization taking place in T cells from CD30L+ AML. The notion that IL-4 was able to enhance in vitro proliferation of CD30L+/IL-4R+ purified leukaemic blasts suggests that the selective interaction of IL-4-producing CD30+ T cells with CD30L+ leukaemic progenitors may have a role in the progression of this particular AML subset.     

Successive Inoculations of Pigs with Porcine Reproductive and Respiratory Syndrome Virus 1 (PRRSV-1) and Swine H1N2 Influenza Virus Suggest a Mutual Interference between the Two Viral Infections

Porcine reproductive and respiratory syndrome virus (PRRSV) and swine influenza A virus (swIAV) are major pathogens of the porcine respiratory disease complex, but little is known on their interaction in super-infected pigs. In this study, we investigated clinical, virological and immunological outcomes of successive infections with PRRSV-1 and H1N2 swIAV. Twenty-four specific pathogen-free piglets were distributed into four groups and inoculated either with PRRSV at study day (SD) 0, or with swIAV at SD8, or with PRRSV and swIAV one week apart at SD0 and SD8, respectively, or mock-inoculated. In PRRSV/swIAV group, the clinical signs usually observed after swIAV infection were attenuated while higher levels of anti-swIAV antibodies were measured in lungs. Concurrently, PRRSV multiplication in lungs was significantly affected by swIAV infection, whereas the cell-mediated immune response specific to PRRSV was detected earlier in blood, as compared to PRRSV group. Moreover, levels of interferon (IFN)-α measured from SD9 in the blood of super-infected pigs were lower than those measured in the swIAV group, but higher than in the PRRSV group at the same time. Correlation analyses suggested an important role of IFN-α in the two-way interference highlighted between both viral infections.     

Expression of MAC-1 (CD11b) in acute myeloid leukemia (AML) is associated with an unfavorable prognosis

There is evidence to suggest, that cellular adhesion molecules and receptors could play a role in leukemia, e.g., through altered adhesive qualities of leukemic blasts. We have studied the expression of the beta2-integrin Mac-1 (CD11b) on mononuclear cells in 48 patients with AML at first diagnosis by flow cytometry using a direct fluorescein-conjugated antibody. A case was defined as positive if more than 20% of the cells expressed Mac-1. Within the FAB types, we observed a high expression rate in cases with M5 (100% MAC-1+ cases, 73% MAC-1+ cells), M4 (75% MAC-1+ cases, 48% MAC-1+ cells) and in cases with FAB-M1 with 71% MAC-1+ cases and 29% MAC-1+ cells. Separating our patients' cohort in cytogenetic risk groups, we could detect significant higher proportions of MAC-1+, cases (88% vs. 27%, P = 0.005) and cells (51% vs. 16%, P = 0.015) with poor cytogenetic risk compared to the favorable risk group. For clinical evaluations only patients treated according to the protocols of the German AML Cooperative Group (AML-CG) were included (n = 29, cases with AML-M3 were excluded). More MAC-1+ cases and cells were found in the "non-responders" group (n = 8) compared to the "responders" group (n = 24). We can conclude that AML cases with high MAC-1 expression are characterized by a worse prognosis. Evaluation of MAC-1 expression in AML might therefore contribute clinically important data with respect to develop new therapies that influence the interactions between integrins like MAC-1 on leukemic cells and endothelial or immunoreactive cells.