Peripheral cellular and humoral responses to infestation with the cattle tick Rhipicephalus microplus in Santa Gertrudis cattle

Resistance to cattle tick infestation in single‐host ticks is primarily manifested against the larval stage and results in the immature tick failing to attach successfully and obtain a meal. This study was conducted to identify immune responses that characterize the tick‐resistant phenotype in cattle. Thirty‐five tick‐naïve Santa Gertrudis heifers were used in this study, thirty of which were artificially infested for thirteen weeks with tick larvae while five animals remained at a tick‐free quarantine property to serve as a control group. Following thirteen weeks of tick infestation, the animals in this trial exhibited highly divergent tick‐resistant phenotypes. Blood samples collected throughout the trial were used to measure peripheral immune parameters: haematology, the percentage of cellular subsets comprising the peripheral blood mononuclear cell (PBMC) population, tick‐specific IgG₁ and IgG₂ antibody titres, IgG1 avidity for tick antigens and the ability of PBMC to recognize and proliferate in response to stimulation with tick antigens in vitro. The tick‐susceptible cattle developed significantly higher tick‐specific IgG₁ antibody titres compared to the tick‐resistant animals. These results suggest that the heightened antibody response either does not play a role in resistance or might contribute to increased susceptibility to infestation.     

Expression of the leucocyte common antigen (LCA,CD45) isoforms RA and RO in acute haematological malignancies: possible relevance in the definition of new overlap points between normal and leukaemic haemopoiesis

Summary. The membrane expression of CD45RA and CD45RO on fresh leukaemic cells taken from 529 cases of acute haemopoietic malignancies, including 117 B-origin acute lymphoblastic leukaemia (B-origin ALL), 3 7 T-origin acute lymphoblastic leukaemia (T-origin ALL), 297 de novo acute myeloid leukaemia (AML), 42 refractory anaemia with excess of blasts in transformation (RAEB-T) and 36 myeloid blastic phase of chronic myelogenous leukaemia (CML-BP-my), was analysed. B-origin ALLs were characterized by the lack of the RO isoform along with the consistent presence of RA. Conversely, a differential expression of the two isoforms was detected in different subsets of T-origin ALL, in that T-stem cell leukaemias (T-SCL: CD7⁺, CD4^?, CD8^?, CD1^?) preferentially expressed CD45RA whereas conventional T-acute lymphoblastic leukaemias (T-ALL: CD7⁺, CD4⁺ and/or CD8⁺ and/or CD1+) were consistently marked by CD45RO. Within myeloid malignancies, most of AMLs displayed CD45RA, while a substantial group of CML-BP-my preferentially exhibited CD45RO. As a general rule, a reciprocal exclusion of the two isoforms was observed in AML as well as in ALL. Nevertheless, a frequent coexpression of CD45RA and CD45RO was observed in CD14⁺ AML. In vitro treatment with all-trans retinoic acid (ATRA) was able to promote a switch from CD45RA to CD45RO expression in 2 7 de novo AML, independently from morphological subtyping. To our knowledge, this is the first report on CD45 isoform expression in a large series of patients with acute leukaemia. The knowledge of the differential expression of CD45RA and CD45RO can ameliorate our classificative approach to haematological malignancies, as well as disclose new multiple overlap points between normal and leukaemic cell differentiation.     

Lymphocyte Subsets in Peripheral Blood and Smoking Habits

The investigation of peripheral blood lymphocyte (PBL) subpopulations is of interest in a wide variety of inflammatory diseases. Since the number of circulating lymphocytes has been shown to be affected by smoking habits, it seems useful to know how PBL subpopulations are influenced. We therefore determined percentages and absolute numbers of a wide range of PBL subpopulations in smokers (n= 14) and nonsmokers (n= 14). PBLs were obtained from healthy volunteers and analyzed by flow cytometry using antibodies for the detection of CD3, CD4, CD8, CD19, CD56, CD57, CD45RO, CD45RA, α/β and γ/δ T cell receptor epitopes. With the exception of CD3⁺ cells, no differences between smokers and nonsmokers were found regarding percentages of PBL subpopulations. Smokers were found to have higher absolute numbers of PBLs in the following subpopulations compared with nonsmokers: CD3⁺, CD4⁺, CD3⁺α/β⁺, CD45RO⁺/CD4⁺, and CD45RA⁺/CD4⁺. Cytotoxic lymphocytes, natural killer cells, and B cells did not differ in number between smokers and nonsmokers. There was likewise no difference in the number of the CD8⁺α/β⁺ and all cells bearing the γ/δ T cell receptor. Smoking increased the number of T cells and mainly CD4⁺ PBLs. The smoking habits of healthy control groups should therefore be taken into account when comparing lymphocyte subpopulations in different diseases. Accepted for publication: 13 February 1997     

Analysis of T cell subsets present in the peripheral blood and synovial fluid of reactive arthritis patients

OBJECTIVE—Reactive arthritis (ReA), a HLA-B27 associated arthropathy, develops in susceptible people after infection with certain bacteria. T cells have been implicated in the pathogenesis of the arthritis but which of the different subsets is involved is still debated. This study has further elucidated the role of the CD4⁺ and CD8⁺ T cells by examining the expression of various surface markers associated with activation.
METHODS—Three colour flow cytometry was used to examine the phenotype of the T cells within the synovial fluid (SF) and peripheral blood (PB) of ReA patients.
RESULTS—ReA SF, compared with paired PB, contained a higher percentage of CD69⁺, CD25⁺, and HLA-DR⁺ CD3⁺ T cells. The majority of SF T cells also expressed the putative memory marker CD45RO. Within the T cell subsets, CD25 was expressed primarily on the CD4⁺ T cells; however more CD8⁺ T cells were HLA-DR⁺.
CONCLUSION—The results show that both CD4⁺ and CD8⁺ T cell populations demonstrate evidence of recent activation. Whether these cells are involved in inducing inflammation, regulating the inflammation, or have become active as a result of migration through the endothelium, remains to be determined by functional studies.

Keywords: reactive arthritis; T cell; peripheral blood; synovial fluid     

Fas/Fas Ligand Expression and Characteristics of Primed CD45RO+ T Cells in the Inflamed Mucosa of Ulcerative Colitis

Background: Chronic immune activation in the colon is characteristic of ulcerative colitis (UC). Fas/Fas ligand (FasL) system is a mechanism responsible for activation-induced cell death (AICD), which maintains homeostasis within the immune system. Thus, Fas/FasL expression on activated colonic T cells of UC patients, as well as the susceptibility of such T cells to AICD was investigated in order to determine the role of activated colonic T cells in the long lasting inflammation in UC. Methods: Fas, FasL, and CD45RO expression on peripheral blood and colonic T cells of UC patients were assayed by flow cytometry. Apoptosis of colonic T cells induced by anti Fas antibody was assessed using the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) assay. Results: The majority of colonic T cells expressed both CD45RO and Fas in the colonic mucosa, a situation that was quite different from that in the peripheral blood. The number of CD45RO⁺CD8⁺ and Fas⁺CD8⁺ T cells was significantly lower in UC patients than the controls, unlike the number of Fas⁺CD4⁺ T cells. In contrast, the number of both CD45RO⁺CD4⁺ and CD45RO⁺CD8⁺ T cells in UC mucosa expressing FasL was significantly higher than in the controls. While Fas mediated apoptosis of CD45RO⁺CD8⁺ T cells was higher in UC patients than the controls, the number of apoptotic CD45RO⁺CD4⁺ T cells from UC mucosa was not. Conclusions: In UC patients, CD45RO⁺CD4⁺ T cells are less sensitive to apoptotic signals mediated by Fas. These phenomena may contribute to the pathogenesis of UC.     

Analysis of iron superoxide dismutase‐encoding DNA vaccine on the evolution of the Leishmania amazonensis experimental infection

The present work aimed to evaluate the immunogenicity of Leishmania amazonensis iron superoxide dismutase (SOD)‐encoding DNA experimental vaccine and the protective properties of this DNA vaccine during infection. The SOD gene was subcloned into the pVAX1 plasmid, and it was used to immunize BALB/c mice. Twenty‐one days after the last immunization, mice were sacrificed (immunogenicity studies) or subcutaneously challenged with L. amazonensis (studies of protection), and alterations in cellular and humoral immune responses were evaluated, as well as the course of infection. Mice only immunized with pVAX1‐SOD presented increased frequencies of CD4⁺IFN‐γ⁺, CD8⁺IFN‐γ⁺ and CD8⁺IL‐4⁺ lymphocytes; moreover, high levels of IgG2a were detected. After challenge, mice that were immunized with pVAX1‐SOD had increased frequencies of the CD4⁺IL‐4⁺, CD8⁺IFN‐γ⁺ and CD8⁺IL‐4⁺ T lymphocytes. In addition, the lymph node cells produced high amounts of IFN‐γ and IL‐4 cytokines. Increased IgG2a was also detected. The pattern of immunity induced by pVAX1‐SOD partially protected the BALB/c mice from a challenge with L. amazonensis, as the animals presented reduced parasitism and lesion size when compared to controls. Taken together, these results indicate that leishmanial SOD modulates the lymphocyte response, and that the elevation in IFN‐γ possibly accounted for the decreased skin parasitism observed in immunized animals.     

Analysis of T-lymphocyte subpopulations in inflammatory bowel diseases by three-color flow cytometry

In order to better define changes in the relative proportion of peripheral blood T-lymphocyte subpopulations in patients with inflammatory diseases of the bowel, we performed simultaneous three-color fluorescence-activated cytometric (FACS) analysis using fluorophore-conjugated monoclonal antibodies with specificity for CD4, CD8, Leu 8, and CD45RA on 22 normal control subjects, 28 patients with Crohn's disease (CD), 15 patients with ulcerative colitis (UC), and 11 patients with intestinal inflammation secondary to etiologies other than inflammatory bowel disease (NIBD). This staining combination allowed enumeration of distinct T-cell subpopulations as follows: virgin CD4⁺, recall antigen helper T cells, nonspecific B-cell helper T cell, virgin CD8⁺, cytotoxic effector and suppressor effector and recall antigen cytotoxic T cells based on a synthesis of published functional analyses. No differences in the proportion of CD4⁺ or CD8⁺ cells or in the CD4⁺/CD8⁺ ratios were evident when UC and NIBD patients were compared to normal subjects. A significant reduction in the proportion of CD4⁺ cells and an increase in CD8⁺ cells was observed, however, in the CD group. When two-color analysis was performed, several significant differences in the proportions of circulating lymphocytes were seen. Specifically, these included significant increases in the number of CD4⁺, Leu 8^– (P<0.01) cells in all disease groups and an increase in CD4⁺, CD45RA⁺ cells in the NIBD group. Conversely, significant decreases in the proportions of CD8⁺, Leu 8⁺ (P<0.01) cells were evident in the Crohn's disease group. Three-color FACS analysis revealed significant differences in the relative proportions of the defined T-cell subpopulations enumerated in the various groups as compared with the normal controls. These included a decrease in the proportions of Leu 8⁺, CD45RA⁺, (virgin) CD8⁺ T cells (P<0.05) and Leu 8^–, CD45RA⁺, (putative recall antigen helper) CD4⁺ T cells (P<0.01) in all patient groups as compared with normal controls. Conversely, an increase in the proportions of Leu 8^–, CD45RA⁺, (putative suppressor effector) CD8⁺ T cells (P<0.01), and Leu 8^–, CD45RA⁺, (function unknown) CD4⁺ T cells (P<0.05) was seen in all patient groups as compared with normal controls. CD patients but not UC or NIBD patients demonstrated a significant increase in the proportion of Leu 8^–, CD45RA⁺, (putative cytotoxic effector) CD8⁺ T cells (P<0.01). An increase in the ratio of Leu 8^–, CD45RA⁺, CD8⁺ (suppressor effector)/Leu 8⁺, CD45RA^–, CD8⁺ (putative cytotoxic effector) T cells was observed in all of the patient groups, but was most accentuated in those with Crohn's disease. Significant decreases in the ratios of Leu 8^–, CD45RA^–, CD4⁺ (putative nonspecific B cell helper)/Leu 8⁺, CD45RA^–, CD4⁺ (recall antigen helper) T cells and Leu 8⁺, CD45RA^–, CD4⁺ (recall antigen helper)/Leu 8^–, CD45RA⁺, CD4⁺ (function unknown) T cells were observed in all of the disease groups studied as compared with normal controls. These results suggest that the proportions of certain peripheral blood T-cell subpopulations are significantly altered in gastrointestinal inflammatory states. Further analysis of these T-lymphocyte subpopulations in the blood and tissues might provide valuable insights into immunological aberrations in inflammatory bowel diseases and might be of value in distinguishing among inflammatory diseases of the intestine.     

Mixed phenotype acute leukemia of T/myeloid type with a prominent cellular heterogeneity and unique karyotypic aberration 45,XY, dic(11;17)

Introduction. A 26‐yr‐old male patient with mixed phenotype acute leukemia of T/myeloid type with prominent leukemic cell heterogeneity, and the presence of a so far unreported karyotype aberration in this type of acute leukemia 45,XY, dic(11;17)(11qter→11p11.2::17p11.2→17qter) is presented. Methods. Flow immunocytometry was performed by direct multicolor immunofluorescent technique on bone marrow aspirates. Cytogenetic analyses were performed using G‐banding method by direct preparation of unstimulated bone marrow cells and following 24 hours of culture in RPMI 1540 culture medium with 25% fetal calf serum at 37°C Results. The flow immunocytometry of bone marrow nucleated cells revealed the existance of three distinct blast cell populations with overlapping immunophenotypes. Predominant blast cell population had an early myeloid phenotype and aberrant expression of CD7 antigen (HLA‐DR⁺, CD34⁺, anti‐MPO⁺, CD117⁺, CD33⁺, CD13⁺, CD7^+low, cyCD3^?, TdT^?). The other two blast cell populations, smaller in cell diameter and less sizable in cell proportion, both shared the T‐lymphoid features. The patient was treated with ADE protocol (etoposide, cytarabine and doxorubicine). A complete remission was achieved and lasted 5 months. Conclusion. A case of MPAL with complex biological features, 45,XY, dic(11;17)(11qter→11p11.2::17p11.2→17qter) karyotype and an aggressive, therapy‐resistant clinical course, is presented.     

Evaluation of T cells in blood after a short gluten challenge for coeliac disease diagnosis

Background and aimTo diagnose coeliac disease (CD) in individuals on a gluten free diet (GFD), we aimed to assess the utility of detecting activated γδ and CD8 T cells expressing gut-homing receptors after a short gluten challenge.MethodsWe studied 15 CD patients and 35 non-CD controls, all exposed to three days of gluten when following a GFD. Peripheral blood was collected before and six days after starting gluten consumption, and the expression of CD103, β7 and CD38 in γδ and CD8 T cells was assessed by flow cytometry. Determination of IFN-γ and IP-10 was performed by means of ELISPOT and/or Luminex technology.ResultsWe observed both γδ and CD8 T cells coexpressing CD103, β7^hi and CD38 in every patient with CD on day six, but only in one control. The studied CD8 T subpopulation was easier to detect than the γδ subpopulation. Increased IFN-γ and IP-10 levels after challenge were observed in patients with CD, but not in controls.ConclusionA short three-day gluten challenge elicits the activation of CD103⁺ β7^hi CD8⁺ T cells in CD. These cells can be detected by flow cytometry in peripheral blood, opening new possibilities for CD diagnosis in individuals on a GFD.     

Selective loss of peripheral blood CD45RO+ T lymphocytes correlates with increased levels of serum cytokines and endothelial cell-derived soluble cell adhesion molecules in patients with chronic idiopathic neutropenia of adults

 The present study was designed to investigate the hypothesis that selective loss of peripheral blood CD45RO⁺ T lymphocytes in patients with chronic idiopathic neutropenia of adults (CINA), previously reported from our laboratory, may be due to enhanced extravasation into the tissues. Serum levels of endothelial cell-derived soluble cell adhesion molecules (sELAM, sICAM and sVCAM), usually used as indicators of endothelial cell activation, were measured in 73 CINA patients and 32 healthy volunteers using a micro-ELISA method. We found that patients had markedly elevated concentrations of all three soluble cell adhesion molecules studied compared to the controls, and serum levels of sELAM, sICAM and, more importantly, sVCAM correlated inversely with the numbers of both CD4⁺/CD45RO⁺ and CD8⁺/CD45RO⁺ T cell subsets. Using a micro-ELISA method, we also measured serum levels of two endothelial cell activators, interleukin (IL)-1β and TNF-α, and found that CINA patients had significantly higher cytokine concentrations than control subjects. Serum levels of IL-1β and TNF-α correlated positively with the values of all three soluble cell adhesion molecules and inversely with the numbers of CD4⁺/CD45RO⁺ and CD8⁺/CD45RO⁺ T cell subsets. Moreover, we measured serum levels of the chemokine RANTES by a micro-ELISA technique and found that CINA patients also had elevated concentrations of the molecule compared to controls. Serum RANTES correlated positively with IL-1β, TNF-α, sICAM, sVCAM and sELAM and inversely with the numbers of both CD4⁺/CD45RO⁺ and CD8⁺/CD45RO⁺ T cell subsets. These findings strongly suggest that CINA patients have an activated endothelium to which CD45RA⁺ and CD45RO⁺ T cells tether and roll, but firm adhesion and transendothelial migration are restricted to CD45RO⁺ T cell subsets, as endothelial VCAM-1 interacts with the vascular leukocyte adhesion molecule-4 (VLA-4) constitutively expressed on CD45RO⁺ but not on CD45RA⁺ T cells. Subsequent subendothelial and tissue migration of CD45RO⁺ T cells may be facilitated by the chemokine RANTES, which acts mainly on CD45RO⁺ T cells. We concluded that selective loss of peripheral blood CD45RO⁺ T lymphocytes in CINA patients is probably due, at least in part, to enhanced extravasation of both CD4⁺/CD45RO⁺ and CD8⁺/CD45RO⁺ T cell subsets into the tissues. Received: February 18, 1998 / Accepted: June 10, 1998     

Altered T-cell subtypes in spondyloarthritis, rheumatoid arthritis and polymyalgia rheumatica

The objective of the present study was to assess the prevalences of naive, memory, memory/effector, regulatory and activated T-cells in peripheral blood (PB) and synovial fluid (SF) of patients with spondyloarthritis (SpA), rheumatoid arthritis (RA), polymyalgia rheumatica/giant cell arteritis (PMR/GCA) and healthy controls (HC). Twenty-two patients with SpA, 15 patients with RA, 38 patients with PMR/GCA and 17 HC were prospectively enrolled. The expression of differentiation and activation markers (CD3, CD4, CD8, CD25, CD28, CD45RA, CD45RO) characterizing T-cell subsets were analyzed by flow cytometry. The frequency of CD3⁺CD4⁺CD28⁻ memory/effector T-cells was increased in PB of patients with SpA (median 1.1%, range 0.1–69.6), RA (2.5%, 0–42.9) and PMR/GCA (2.7%, 0–49.5) when compared with HC (0.7%, 0–38.0) and tended to be higher in SF of SpA patients (4.5%, 0.2–7.2, P = 0.084). CD28⁺CD45RA⁺CD4⁺ (9.6%, 4.1–10.3) and CD28⁺CD45RA⁺CD8⁺ naive T-cells (15.0%, 12.9–26.2) were reduced and CD28⁺CD45RO⁺CD4⁺ (93.5%, 51.0–99.0), CD28⁺CD45RO⁺CD8⁺ memory (81.2%, 38.9–83.5), CD8⁺CD25⁺ activated T-cells (10.9%, 2.7–13.8) and CD4⁺CD25^hi TREGs (10.2%, 7.0–13.3) were increased in SF compared to PB (P < 0.05 each). These findings demonstrate altered T-cell subsets in patients with immune-mediated disease, particularly at sites of inflammation.     

Lymphotropic HCV strain can infect human primary na?ve CD4+ cells and affect their proliferation and IFN-�� secretion activity

Background

Lymphotropic hepatitis C virus (HCV) infection of B and T cells might play an important role in the pathogenesis of hepatitis C. Recently, we showed that a lymphotropic HCV (SB strain) could infect established T-cell lines and B-cell lines. However, whether HCV replication interferes with cell proliferation and function in primary T lymphocytes is still unclear.

Aim

The aim of this study was to analyze whether HCV replication in primary T lymphocytes affected their development, proliferation, and Th1 commitment.

Methods

SB strain cell culture supernatant (2?×?10⁴?copies/ml HCV) was used to infect several kinds of primary lymphocyte subsets. Mock, UV-irradiated SB-HCV, JFH-1 strain, and JFH-1 NS5B mutant, which could not replicate in T cells, were included as negative controls. Carboxyfluorescein succinimidyl ester (CFSE) and CD45RA double staining was used to evaluate the proliferative activity of CD4⁺CD45RA⁺CD45RO^? na?ve CD4⁺ cells. Interferon (IFN)-?? and interleukin (IL)-10 secretion assays magnetic cell sorting (MACS) were carried out.

Results

Negative strand HCV RNA was detected in CD4⁺, CD14⁺, and CD19⁺ cells. Among CD4⁺ cells, CD4⁺CD45RA⁺RO^? cells (na?ve CD4⁺ cells) were most susceptible to replication of the SB strain. The levels of CFSE and CD45RA expression gradually declined during cell division in uninfected cells, while HCV-infected na?ve CD4⁺ cells expressed higher levels of CFSE and CD45RA than Mock or UV-SB infected na?ve CD4⁺ cells. Moreover, the production of IFN-?? was significantly suppressed in SB-infected na?ve CD4⁺ cells.

Conclusions

Lymphotropic HCV replication suppressed proliferation and development, including that towards Th1 commitment, in human primary na?ve CD4⁺ cells.     

Melatonin: Antioxidant and modulatory properties in age‐related changes during Trypanosoma cruzi infection

The purpose of this study was to investigate the effects of melatonin on selected biomarkers of innate and humoral immune response as well as the antioxidant/oxidant status (superoxide dismutase—SOD and reduced glutathione levels (GSH) to understand whether age‐related changes would influence the development of acute Trypanosoma cruzi (T. cruzi) infection. Young‐ (5 weeks) and middle‐aged (18 months) Wistar rats were orally treated with melatonin (gavage) (05 mg/kg/day), 9 days after infection. A significant increase in both SOD activity and GSH levels was found in plasma from all middle‐aged melatonin‐treated animals. Melatonin triggered enhanced expression of major histocompatibility class II (MHC‐II) antigens on antigen‐presenting cell (APC) and peritoneal macrophages in all treated animals. High levels of CD4⁺CD28‐negative T cells (*P<.05) were detected in middle‐aged control animals. Melatonin induced a significant reduction (***P<.001) in CD28‐negative in CD4⁺ and CD8⁺ T cells in middle‐aged control animals. Contrarily, the same group displayed upregulated CD4⁺CD28⁺T and CD8⁺CD28⁺T cells. Melatonin also triggered an upregulation of CD80 and CD86 expression in all young‐treated groups. Significant percentages of B and spleen dendritic cells in middle‐aged infected and treated animals were observed. Our data reveal new features of melatonin action in inhibiting membrane lipid peroxidation, through the reduction in 8‐isoprostane, upregulating the antioxidant defenses and triggering an effective balance in the antioxidant/oxidant status during acute infection. The ability of melatonin to counteract the immune alterations induced by aging added further support to its use as a potential therapeutic target not only for T. cruzi infection but also for other immunocompromised states.     

Acquired resistance to ticks: expression of resistance by C4-deficient guinea pigs.

Hartley and C4-deficient guinea pigs developed resistance to the ixodid tick. Dermacentor andersoni, after one infestation. Resistance was characterized by resistant animals of both groups allowing significantly fewer larvae (5--25%) to engorge during a second infestation than during an initial infestation (70--90%). Resistant animals in both groups developed cutaneous reactions at the site of tick attachment which were characterized by intraepidermal vesicles containing numerous basophils. In previous studies, tick-resistant Hartley guinea pigs depleted of complement by cobra venom factor were not able to express the resistance response and the skin reactions at the tick attachment sites were depleted of basophils. The use of cobra venom factor as an anti-complement probe could not distinguish the relative importance of the classical and/or alternate pathways of complement activation in the expression of tick resistance. The present study reports that C4-deficient guinea pigs, those with a total deficiency in the classical pathway of complement activation, but with an intact alternate pathway, can acquire and display tick resistance in a fashion similar to Hartley guinea pigs. This finding provides evidence that the alternate pathway of complement activation is important in the expression of tick resistance.     

The role of viruses in Type I diabetes: two distinct cellular and molecular pathogenic mechanisms of virus-induced diabetes in animals

Type I (insulin-dependent) diabetes mellitus results from the progressive loss of pancreatic beta cells. Environmental factors are believed to play an important part in the development of Type I diabetes by influencing the penetrance of diabetes susceptibility genes. As one environmental factor, the virus has long been considered to play a part in this disease. To date 13 different viruses have been reported to be associated with the development of Type I diabetes in humans and in various animal models. The most clear and unequivocal evidence that a virus induces diabetes in animals comes from studies on the d variant of the encephalomyocarditis (EMC-D) virus in mice and the Kilham rat virus (KRV) in rats. The infection of genetically susceptible strains of mice with a high titre of EMC-D virus results in the development of diabetes within 3 days. This is largely due to the rapid destruction of beta cells by the replication of the virus within the beta cells. In contrast, the infection of mice with a low titre of EMC-D virus results in a limited replication of the virus before the induction of neutralizing anti-virus antibody and the subsequent recruitment of activated macrophages. The Src kinases, particularly hck, play an important part in the activation of macrophages and the subsequent production of tumour necrosis factor (TNF)-α, interleukin (IL)-1β and nitric oxide (NO), leading to the destruction of beta cells which results in the development of diabetes. The Kilham rat virus causes autoimmune diabetes in diabetes resistant (DR)-BB rats without infection of beta cells. The infection of DR-BB rats with KRV results in the disruption of the finely tuned immune balance of Th1-like CD45RC⁺CD4⁺ and Th2-like CD45RC^-CD4⁺ T cells, leading to the selective activation of beta-cell-cytotoxic effector T cells. [Diabetologia (2001) 44: 271–285]     

Mucosal tolerance of the hookworm Ancylostoma caninum in the gut of naturally infected wild dogs

Ancylostoma caninum is a very pathogenic hookworm that locates in the small intestine of the dog and other canid species. The mucosal response of wild dogs naturally infected with A. caninum was investigated in this study. In spite of diffuse infiltrations of the mucosa with CD3⁺, CD4⁺, CD8⁺, CD11c⁺, CD21⁺ or MHC class II antigen cells, no focal infiltrations with any of these cell phenotypes were observed around the buccal capsule or the body of the feeding worms. Very few or no apoptotic cells could be detected around the worms fixed into the mucosa but they were detected on the tip of villi and in the superficial layer of cellular debris and proteinaceous exudate that covers the mucosa. Muc5AC, a mucine associated with expulsion of gut worms (Trichuris muris) was expressed extremely weakly or was not expressed at all in the intestine of the wild dogs infected with A. caninum. Our data show that individual specimens of A. caninum can reside for some time in the mucosa of the gut of dogs undetected and most likely unaffected by the effectors of the local immune response.     

Circulating Endothelial Progenitor Cells as Markers for Severity of Ischemic Chronic Heart Failure

IntroductionDespite a high potential of endothelial progenitor cells (EPCs) for diagnostic purposes, the EPC role in developing ischemic chronic heart failure (CHF) has not been determined obviously.The objective of this study was to assess the counts of CD45⁺CD34⁺, CD45⁻CD34⁺, CD14⁺CD309⁺, and CD14⁺CD309⁺Tie2⁺ phenotyped circulating EPCs of various subpopulations in patients with ischemic CHF.Methods and ResultsThe study involved 153 patients (86 male), aged 48–62 years, with angiographically proven coronary artery disease (CAD) and 25 healthy volunteers. CHF was diagnosed in 109 patients (71.2%). Mononuclear cell populations were phenotyped by flow cytofluorimetry. Cardiovascular risk factors, such as type 2 diabetes mellitus, hyperlipidemia, arterial hypertension, and adherence to smoking, may have a negative effect on circulating EPC counts in CAD patients regardless of the presence of CHF. The depletion of the CD14⁺CD309⁺- and CD14⁺CD309⁺Tie2⁺-phenotyped circulating EPC counts is associated with the severity of left ventricular dysfunction, whereas the CD45⁺CD34⁺- and CD45⁻CD34⁺-mononuclear cell counts are more representative of the severity of atherosclerotic coronary artery lesions.ConclusionThe authors found that New York Heart Association functional class of CHF, left ventricular ejection fraction <42%, the N-terminal pro–B-type natriuretic peptide level >554 pg/mL, and Е/Еm ratio >15 U had the highest predictive value for the depletion of the EPC count in CAD patients.     

Associations of circulating CXCR3–PD-1+CD4+T cells with disease activity of systemic lupus erythematosus

Abstract

Objectives: Which helper CD4⁺ T cell subset contributes to autoantibodies generation and severity of end-organ involvement in lupus patients remains to be explored. Our research aims to investigate the roles of circulating Tfh (cTfh) cell subsets and corresponding CXCR5^– Th cells in lupus patients and their correlation with SLE disease activity index 2000 (SLEDAI).

Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from blood of systemic lupus erythematosus (SLE) patients as well as healthy donors. The proportion of Th cell subsets classified from cell surface markers (CD45RO, CXCR5, CXCR3, CCR6, PD-1, ICOS, and CCR7) is detected by flow cytometry.

Results: We found no difference in the frequency of CD45RO⁺CXCR5⁺CD4⁺ T cells between SLE patients and health controls. As previously reported, SLE patients showed an increase in the percentage of CXCR5⁺PD-1⁺, CXCR5⁺ICOS⁺PD-1⁺ and CXCR5⁺CCR7^loPD-1^hi cTfh subset, however, none of these populations had correlation with SLEDAI. Therefore, we further investigated the CXCR5^– subsets, and surprisingly we found that the frequency of CXCR3^–PD-1⁺ subset was correlated with SLEDAI, ds-DNA IgG, anti-nucleosome antibody, C3, and C4 independent of CXCR5. Consistently, CXCR3^–PD-1⁺CD45RO⁺CD4⁺T cells expressed factors associated with B-cell-help for the autoantibody production.

Conclusion: CXCR3^–PD-1⁺CD4⁺T cells are a sensitive indicator to assess SLE disease activity and might contribute B cell help and the generation of autoantibodies in patients.     

Detection of Bm86 antigen in different strains of Boophilus microplus and effectiveness of immunization with recombinant Bm86

The control of tick populations by using conventional strategies poses several problems, including the appearance of organophosphate resistant strains, among others. The possibility of using alternative strategies such as vaccination with tick antigens has been suggested by several authors. One particular antigen (Bm86) has been described and shown to be able to induce a protective immunity against the cattle tick Boophilus microplus. In this paper we demonstrate by means of immunohistochemical staining that this antigen is conserved among several strains of this species. These results correlate with those showing that animals vaccinated with a preparation of recombinant Bm86 were protected against challenge with the four different strains tested, including one resistant to organophosphates. These results favour the immunization with recombinant Bm86 for the control of the cattle tick B. microplus.