bluuebunny/crossref_metadata_2025 huggingface dataset:
Analysis of thein vitrocytokine production by liver-infiltrating T cells of patients with autoimmune hepatitis
Authors: SCHLAAK J F , LÖHR H , GALLATI H , MEYER ZUM BÜSCHENFELDE K-H , FLEISCHER B
Published: July 2010

The pathogenic mechanisms underlying the development of autoimmune hepatitis (AIH) are still unclear. Since AIH is associated with the presence of various autoantibodics and certain HLA subtypes, it is likely that T and B cells play a major role in this disease. In this study we have determined the functional capacities of in vivo preactivated liver-infiltrating T cells (LTC) from patients with AIH. As controls we used LTC from patients with non-autoimmune hepatitis (non-AIH). Our results show that preactivated LTC from patients with AIH predominantly (190/255 clones) reside in the CD4+ population, whereas LTC in non-AIH are dominated by the CD8+ phenotype (148/254 clones). In view of this finding we have investigated the cytokine secretion patterns of 102 randomly chosen CD4+ T cell clones from six patients with AIH. As controls we have used 58 CD4+ LTC from 11 patients with non-AIH. All clones were stimulated by lectin and irradiated accessory cells and subsequent cytokine production was evaluated. LTC from patients with AIH have a lower interferon-gamma (IFN-γ)/IL-4 ratio compared with LTC from non-AIH. Although clones from some patients with AIH produced very high amounts of IL-4 in vitro, this was not a constant finding. These results show that in two preactivated LTC from patients with AIH are mostly CD4+ T cells that produce more IL-4 than IFN-γ. In contrast. LTC from patients with non-AIH are dominated by CD8+ and CD4+ T cells that produce significantly less IL-4 than IFN-γ. Thus, liver-infiltrating T cells from patients with AIH and non-AIH belong to different functional T cell subsets. This may have implications for the regulation of humoral and cellular immune responses in inflammatory liver disease.
The accumulation of inflammatory cells in synovial sheath and epitenon during adhesion formation in healing rat flexor tendons
Authors: WOJCIAK B , CROSSAN J F
Published: July 2010

The accumulation of inflammatory cells in synovial tissue was studied using indirect immunofluorescence assays on cell cultures and frozen tissue sections of healing rat digital flexor tendons, Flexor tendons were collected from rats 3, 7 and 14 days after crush injury. Tendon sheath and epithenon cells were isolated by sequential enzymic digestion and cultured for 2 days. Subpopulations of synovial and inflammatory cells were identified with MoAbs against cell surface glycoproteins present on B lymphocytes (CD45), T lymphocytes (CD2, CD4, CD8), macrophages (CD14) and endothelial cells. A phagocytosis assay was also used to identify macrophages. We report a substantial increase in the number of T lymphocytes (mainly helper inducer) and phagocytotic cells with monocyte/macrophage surface markers in tendon sheath and epitenon 3 days after crush injury. The infiltration of inflammatory cells into synovial sheath and epitenon preceded an increase in fibronectin production by tendon cells which was seen 7 days after injury. To study the interaction between T lymphocytes and synovial cells in vitro, we established synovial fibroblast-like type B cell cultures and used stimulated and non-stimulated T lymphocytes in cell binding assays. We observed increased adhesiveness between unstimulated synovial cells and synovial cells previously cultured with activated and non-activated T lymphocytes. ELISA inhibition studies have shown an increase in fibronectin production by synovial fibroblastsco-cultured with stimulated CD4+ T lymphocytes. We suggest that the presence of inflammatory cells in synovial sheath and epitenon during tendon healing induces synovial fibroblasts and epilenon cells to increase their production of fibronectin. Which provides a scaffold for subsequent adhesion formation.
Identification of a 46-kD latex protein allergen in health care workers
Authors: BEEZHOLD D H , SUSSMAN G L , KOSTYAL D A , CHANG N-S
Published: July 2010

Latex allergy is an occupational hazard for health care workers. Extractable latex proteins are known to be allergenic, but most latex allergens have not been specifically identified. The purpose of this study was to characterize the IgE response of latex-allergic patients to latex proteins and to identify common protein allergens. Serum was obtained from 40 individuals who were skin test-positive to latex; 85% were health care workers. Western blots for IgE reactivity were performed using both ammoniated (AL) and non-ammoniated (NAL) latex proteins and IgE-reactive NAL proteins were analysed by microsequence analysis. The patients were grouped according to common patterns of reactivity. Pattern 1, the most common pattern of reactivity (9/40 patients) recognized two protein bands in both NAL and AL at 46 and 110kD. A second, heterogeneous pattern of reactivity (pattern 2) recognized a diffuse pattern of polypeptides in the AL preparation. The n-terminal amino acid sequences for allergens at 14, 18, 29, 46 and 110 kD were determined. Sequence analysis identified the 14-kD and 18-kD allergens as the hevein preprotein. The 46-kD and 110-kD had identical sequences which were unique from known latex proteins. We conclude that multiple latex proteins are allergens with hevein preprotein and a previously unidentified 46/110-kD protein being commonly recognized in health care workers.
Human intestinal alkaline phosphatase-binding IgG in patients with severe bacterial infections
Authors: MÄDER M , KOLBUS N , MEIHORST D , KÖHN A , BEUCHE W , FELGENHAUER K
Published: July 2010

Patterns of alkaline phosphatase (AP)-binding proteins were observed in the alkaline pH range of 6-5-9-5 upon isoelectric focusing and blotting of serum from patients with inflammatory diseases. After isolation using affinity chromatography on protein A or immunoaffinity chromatography on AP coupled to cyanogen bromide (CNBr)-activated Sepharose, the AP-binding protein was identified as IgG on Western blots and in ELISA using human IgG-specific antibodies. It was shown that this IgG binds to AP from both calf (bovine) and human intestine. However, it binds neither lo the human liver-bone-kidney (LBK) isoform nor to bacterial AP. Moderate reaction was observed with human placental AP. Comparing patients with various diagnoses (n = 284), AP-binding antibodies were mainly found in severe bacterial infections. They were not detected in serum from healthy blood donors (n = 300). The presence of AP-binding IgG was independent of the infected organ and the bacterial species causing infection. This antibody may be useful for discriminating bacterial from viral infection and for indicating severe bacterial inflammation.
Dichotomy of the human T cell response toLeishmaniaantigens. II. Absent or Th2-like response to gp63 and Thl-like response to lipophosphoglycan- associated protein in cells from cured visceral leishmaniasis patients
Authors: KURTZHALS J A L , HEY A S , JARDIM A , KEMP M , SCHAEFER K-U , ODERA E O , CHRISTENSEN C B V , GITHURE J I , OLAFSON R W , THEANDER T G , KHARAZMI A
Published: July 2010

The T cell response to different Leishmania donovani aniigens was investigated using peripheral blood mononuciear cells (PBMC) from Kenyans cured of visceral leishmaniasis and non-exposed Danes. Crude promastigote and amastigote antigens both induced proliferation and interferon-gamma (IFN-γ) production in PBMC from cured patients, while cells from non-exposed donors gave weak responses. A similar pattern was indticed by lipophosphoglycan-associated protein (LPGAP). By contrast, the major surface protease of Leishmania. gp63, induced only a weak proliferative response without IFN-γ production in five of 17 samples from cured patients. Four of the five responding cultures produced IL-4, i.e. the response to this antigen was of the Th2 type. Furthermore, sera from acutely ill visceral leishmaniasis patients contained high levels of IgG antibodies to gp63. The Th2-tike response to gp63 in patients cured of visceral leishmaniasis differs from the Thl-like response to the same antigen observed in patients cured of cutaneous leishmaniasis.
Flow cytometric analysis of the stimulatory response of T cell subsets from normal and HIV-1+ individuals to various mitogenic stimuliin vitro
Authors: MEDINA E , BORTHWICK N , JOHNSON M A , MILLER S , BOFILL M
Published: July 2010

A novel technique is described which allows the study of the responses of T cell subpopulations stimulated in bulk cultures without interfering with cell-cell interactions. The number and phenotype of lymphoblasts developing following stimulation with phytohaemagglutinin (PHA), anti-CD3, staphylococcal protein A (SPA) and pokeweed mitogen (PWM) was determined in HIV-P and HIV-1+ patients using a new five-parameter flow cytometric method. We found that normal T ceils responded faster to PHA than lo any of the other mitogens tested. The peak of the PHA response occurred on day 3. followed by anti-CD3 and SPA on day 4 and PWM mitogen on day 5. Although PHA and anti-CD3 stimulated up to 95% and 80% of lymphocytes, respectively, SPA and PWM stimulated only 40% and 30% of cells, respectively. A defective T cell response was observed in lymphocytes cultured from asymptomatic HlV-1+ patients compared with negative controls. This loss of response was related to a selective mortality of T cells following mitogenic stimulation, referred to as activation-associated lymphocyte death (AALD). The results showed that stronger mitogens (PHA and anti-CD3) induced AALD in a larger proportion (50–60%) of T cells than weaker mitogens such as SPA and PWM (30–40%). and that AALD affected different lymphocyte subsets to different extents. AALD occurred more frequently in total CD4+ and CD45RO+ T ceils compared with CD4+ and CD45RA+ T cells, but memory CD4+ T cells were the population most severely affected in samples from HIV-I+ donors.
Primary T lymphocyte immunodeficiency associated with a selective impairment of CD2, CD3, CD43 (but not CD28)-mediated signal transduction
Authors: RODRIGUEZ-GALLEGO C , ARNAIZ-VILLENA A , CORELL A , MANZANARES J , TIMON M , PACHHCO A , REGUEIRO J R
Published: July 2010

A 2-year-old female with important signs of immune response failure against virus, bacteria, fungi and protozoa and no obvious humoral or lymphocyte phenotypical defect was studied. Both peripheral blood mononuclear cells and IL-2-dependent T cell lines derived from the patient showed a severe selective T cell activation impairment via CD2, CD3 and CD43; however, this defect was reversible with the addition of either IL-2, or phorbol myristate acetate (PMA) or anti-CD28 antibodies, Concordantly, the induction of IL-2 (and, in part, IL-3 and IL-4) messenger RNA Vk-as severely reduced in stimulated T cells, but that of other cytokines was either normal (IL-5) or only slightly diminished (interferon-gamma (IFN–γ)). It is concluded that an activation T cell defect exists previous to protein kinase C (PKC) and between membrane receptors and the activation pathway of certain response genes encoding for interleukins involved in proliferation (i.e. lL-2, IL-3 and IL-4), but not of others (i.e. IL-5). The use of T ceil lines from human T lymphocyte activation deficiencies allows dissection of T cell pathology and the corresponding physiological pathways. In the present description, there is an evident independence of the CD28 T cell activation pathway from those induced through CD2 or CD3, and the differential gene regulation of the different interleukins.
Flow cytometry CD4+/CD8+ ratio of liver-derived lymphocytes correlates with viral replication in chronic hepatitis B
Authors: PHAM B-N , MOSNIER J F , WALKER F , NJAPOUM C , BOUGY F , DEGOTT C , ERLINGER S , COHEN J H M , DEGOS F
Published: July 2010

T lymphocytes have been assumed to play an essential role in tissue injury in patients with chronic hepatitis B. As hepatitis B virus (HBV) is considered as a major factor controlling liver inflammation, we assessed whether a particular T lymphocyte subset could be preferentially detected in the liver in accordance with viral replication. Liver-derived lymphocytes and peripheral blood lymphocytes were analysed by flow cytometry in 21 patients with histologically confirmed chronic hepatitis B without cirrhosis. Viral replication was quantified by hybridization of serum HBV DNA. Eleven patients exhibited an active viral replication with serum HBV DNA ranging from 10 to 388pg/ml at the time of the liver biopsy, whereas 10 patients had no detectable serum HBV DNA. In patients exhibiting viral replication. CD4+/CD8+ ratios of liver-derived lymphocytes were significantly higher [P < 005) than those obtained in patients without viral replication. In contrast, the percentage of T cells expressing the γ/δ receptor and that of CD2±/CD57+ cells were similar in both groups of patients. Furthermore, in patients exhibiting viral replication. CD4+/CD8+ ratios of liver-derived lymphocytes correlated with serum HBV DNA levels (P<0.0001). No relationship between CD4+/CD8+ ratio of liver-derived and peripheral blood lymphocytes was observed. Our data indicate that, in patients with chronic hepatitis B. the CD4+/CD8+ ratio of liver-derived lymphocytes correlates with viral replication. This suggests that in situ helper/inducer CD4+ T lymphocytes may positively regulate the cytoxic T cell activity in patients with HBV-related chronic hepatitis.
B7/BB1 provides an important costimulatory signal for CD3-mediated T lymphocyte proliferation in patients with systemic lupus erythematosus (SLE)
Authors: SFIKAKIS P P , OGLESBY R , SFIKAKIS P , TSOKOS G C
Published: July 2010

Successful T cell activation via the T cell receptor (TCR)/CD3 complex requires at least one contact-dependent second signal delivered by costimulatory molecules, including the B7/BB1 molecule, that are present on antigen-presenting cells (APC). SLE is characterized by multiple complex lymphocyte abnormalities of undefined molecular origin. It is currently unclear whether an intrinsic defect of T cell or an underlying APC dysfunction is responsible for defective in vitro proliferation of T cells from patients with SLE. We planned the present experiments to ask whether the TCR/CD3-mediated and B7/BB1-costimulated T cell proliferation is normal in these patients. We used enriched T cell populations that were stimulated with an anti-CD3 MoAb in the presence of controlled quantities of functional B7/BB1 antigen. Freshly isolated T cells from 17 SLE patients (10 and seven patients with either active or inactive disease, respectively) and 11 normal individuals were cocultured with irradiated B7/BB1-transfected P815 cells or parental P815 cells in the presence of OKT3 MoAb at optimal and suboptimal concentrations for 2·5-7 days. Normal or SLE T cells responded similarly to stimulation via anti-CD3, in the absence of B7/BB1 antigen. A several-fold increase in T cell proliferation in the presence of B7/BB1 antigen was observed. Proliferation was inhibited in the presence of anti-B7/BB1 MoAb, but not with control MoAbs. Interestingly, dose-response curves and time kinetics of B7/BB1 costimulation were similar in T cells from patients with either active or inactive SLE at the time of study, and normal individuals. In addition, no differences in the IL-2 receptor release by T cells cultured under these conditions were observed between SLE patients and normal individuals. These results demonstrate that CD28 signalling is not intrinsically impaired in patients with SLE; further studies to investigate whether abnormal B7/BB1 expression is involved in the autoimmune process are needed.
T cell reactivity against antigen 85 but not against the 18- and 65-kD heat shock proteins in the early stages of acquired immunity againstMycobacterium leprae
Authors: LAUNOIS P , N'DIAYE M NIANG , SARTHOU J L , DROWART A , VAN VOOREN J P , CARTEL J L , HUYGEN K
Published: July 2010

T cell proliferation and interferon-gamma (IFN-γ) production of peripheral blood mononuclear cells (PBMC) from 20 household contacts were tested against the 18- and 65-kD heat shock proteins from Mycobacterium leprae (ML 18 and ML65 respectively) and antigen 85 from Myco. bovis bacille Calmette-Guerin (BCG) (Ag 85) during a 12-months follow-up study. Among the eight contacts that became positive, eight showed positive reactivity against Ag 85, 5/8 against ML65 and 4/8 against ML 18 at the end of the study. Of the 16 contacts who were lepromin-positive either at first or second testing, all responded to Ag 85, 11 to ML 65, but only eight reacted to ML18 antigen. Contacts who were lepromin-positive at first testing developed responses to ML18 only at second testing. In contrast, among the four contacts that remained lepromin-negative during the follow up, three proliferated to Ag 85 either at first or second testing, but only one produced IFN-γ against Ag 85 at the end of the study. These results demonstrated that T cell reactivity and particularly IFN-γ secretion against Ag 85, but not against ML18 and ML65, might be a predominant mechanism in the early stages of acquired protective immunity against Myco. leprae.
Oral and aerosol immunization with viable or inactivatedActinobacillus pleuropneumoniaebacteria: antibody response to capsular polysaccharides in bronchoalveolar lavage fluids (BALF) and sera of pigs
Authors: HENSEL A , PABST R , BUNKA S , PETZOLDT K
Published: July 2010

To investigate the antibody response after local application of lung-pathogenic bacteria, pigs were immunized with viable or inactivated Actinohacillus pleuropneumoniae by the oral and aerogenous route. After 3 weeks class-specific immunoglobulins against purified A. pleuropneumoniae capsular polysaccharides (CP) were determined in scrum and BALF by ELISA. A significant increase of IgA antibodies was found in BALF but not in sera of all iminunized pigs. Oral immunization with viable A. pleuropneumoniae and aerosol immunization with either viable or inactivated bacteria resulted in a significant increase of IgG antibodies to the CP antigen in BALF, whereas only aerosol exposure to viable bacteria resulted in a significant increase in IgG antibodies in serum. A significant increase in anti-CP IgM in BALF was observed after aerosol exposure but not after oral immunization. IgM antibodies towards CP increased significantly by both routes of immunization with viable bacteria. The anti-CP activity of all three isotypes in sera and BALF was low in all groups compared with the positive controls, although inoculation of viable A. pleuropneumoniae led to higher levels of antibody concentration than inactivated bacteria. Our results indicate a traffic of primed lymphocytes from the gut into the bronchoalveolar airways and further support the hypothesis that polysaccharide-specific B cells may functionally mature at the mucosal surfaces.
Increased plasma levels of soluble IL-2R are associated with severe Plasmodium falciparum malaria
Authors: JAKOBSEN P H , MORRIS-JONES S , THEANDER T G , HVIID L , HANSEN M B , BENDTZEN K , RIDLEY R G , GREENWOOD B M
Published: July 2010

Plasma samples from children with mild and severe Plasmodium falciparum malaria and from children with unrelated diseases were collected to investigate whether the clinical outcome of infection was associated with plasma factors which reflected the activity of different cells of the immune system. Children with severe P. falciparum malaria had significantly higher plasma levels of soluble IL-2R than children with mild malaria. Plasma levels of 1L-2R and levels of parasitaemia were significantly correlated. Neither parasitaemia nor plasma levels of tumour necrosis factor-alpha (TNF -a). IL-6. lymphotoxin (LT). interferon-gamma (IFN-γ). IL-4, soluble IL-4R or soluble CD8 differed significantly between the two groups of children with malaria. High plasma levels of soluble CD8 were associated with failure of lymphocytes to produce I FN-γin vitro following stimulation with P. falciparum antigen. We conclude that soluble IL-2R is a useful marker of disease severity independently of the association with levels of parasitaemia, and that functional regulation of different lymphocyte subsets occurs during acute malaria episodes.
Follow up of soluble IL-2 receptor level in metastatic malignant melanoma patients treated by chemoimmunotherapy
Authors: SOUBRANE C L , MOUAWAD R , ICHEN M , SUISSA J , BOREL C H , VUILLEMIN E , BENHAMMOUDA A , BIZZARI J P , WEIL M , KHAYAT D
Published: July 2010

Immunological parameters following chemoimmunotherapy combination were studied in 31 patients with metastatic malignant melanoina. They received Cisplatni (100 mg/m2) on day 1 and 28, recombinant IL-2 (rIL-2; Eurocetus) in continuous infusion from day 3 to 6, 17 to 21, 31 to 34 and 45 to 49. Interferon-alpha (IFN-α: Roche) was given subcutaneously three times weekly. No significant change in CD4/CD8 ratio at onset or during treatment was observed between responder (n=19) and non-responder (n=12) patients. Regarding the IL-2 receptor (IL-2R) study, the percent age of cells expressing Tac(p55)receptor did not change either for healthy volunteers (n=20) and patients before any therapy, or between responder and non-responder patients. Concerning serum soluble IL-2R shedding before therapy. we observed A significant increase (P=0·001) in patients (79±40 pM) compared with healthy donors (30±15 pM), but no significant variation was seen between responder and non-responder patients. In contrast, during the treatment, the soluble IL-2R level increased in both groups but, interestingly, a significant difference was found between responder and non-responder patients from day 7 (P<0·05) to day 21 (P<0·01), suggesting that the cells from non-responder may be slower in becoming stimulated. This finding is the most striking point of our study and suggests that sIL-2R might be an early predictive factor of the clinical response as obtained by logistic regression (P=0·0063). Therefore patients with a serum soluble IL-2R level greater than 250 pM at day 21 have a 12-fold more chance of undergoing a clinical response.
Expression patterns of Fcγ receptors, HLA-DR and selected adhesion molecules on monocytes from normal and HIV-infected individuals
Authors: Locher CH , Vanham G , Kestens L , Kruger M , Ceuppens J L , Vingerhoets J , Gigase P
Published: July 2010

The expression and co-expression profiles of functionally important monocyte surface markers were compared between control and HIV+ individuals using combined physical gating and dim CD4 expression to delineate the monocytes. The FcγRII (CD32), the MHC class II antigen HLADR and the adhesion molecules CDIIa (LFA-1α), CD18 and CD54 (ICAM-1) showed an unimodal distribution. Of these markers, CDI Ia and HLA-DR were up-regulated in the HIV+ subjects compared with controls. The expression levels of the adhesion molecules correlated with each other in both patients and controls. The CD11b (CR3-α), CD14, FcγRI, and FcγRIII markers were bimodally distributed. Compared with controls, monocytes from seropositives contained fewer CDbright+ cells, an equal proportion of FcγIbright+ cells, but twice as many FCγ RIII+ cells. The expression level of FcγRI and CD11b within their brightly positive subset increased as CD4T cells decreased. Both in patients and controls, co-expression of bright CD11b, CD14 and FCγRI was shown, whereas the FcγRIII+ cells were negative or dim positive for the former triad. We conclude that the expression of two FcγR (I and III), of the adhesion molecules CD11a and CD11b and of HLA-DR showed particular alterations on monocytes from HIV+ subjects. The relationship of these phenotypic observations with altered cytokine profiles and altered monocyte function is discussed.
Cytokine production in varicella zoster virus-stimulated limiting dilution lymphocyte cultures
Authors: ZHANG Y , COSYNS M , LEVIN M J , HAYWARD A R
Published: July 2010

Human blood lymphocytes were stimulated with varicella zoster virus (VZV) antigen in limiting dilution cultures and the amounts of interferon-gamma (IFN-γ) and IL-4 measured in the supernatants. The results indicate that up to 85% of proliferating cells of young adults produce IFN-γ and up to 10% make IL-4. At limiting dilution, few if any wells were positive for both IFN-γ and IL-4. The amount of IFN-γ per well increased in the presence of antibody to IL-4, but anti-IFN-γ did not increase IL-4 production. The frequency of wells containing IFN-γ was lower in subjects < 19 or > 55 years of age, and the amounts of IFN-γ in positive wells was significantly lower in cultures of the older subjects' lymphocytes. The frequency of IL-4-making cells did not fall significantly with age. The data suggest that the age-related decline in the frequency of blood T cells which responds to VZV affects mainly the cells with a Th1-like cytokine phenotype.
Immunological markers of childhood fevers in an area of intense and perennial malaria transmission
Authors: HURT N , THEIN M , SMITH T , BORDMANN G , GALLATI H , DREES N , TANNER M , WEISS N
Published: July 2010

In order to describe presumed paediatric malaria on a cell-immunological basis, the soluble receptors of IL-2 (sIL-2R) and tumour necrosis factor (sTNF-R55 and sTNF-R75) were quantified in highly exposed young Tanzanian children. Sera were obtained from 66 acute and 72 reported febrile patients during health post consultations and follow-ups and from 68 community controls. Levels of sIL-2R, sTNF-R55 and sTNF-R75 were significantly elevated during fever attacks, especially in very young children. Soluble TNF-R75 levels were most stable and those of sTNF-R55 least. Levels of sTNF-R55 were related to the magnitude of fever and thus appeared to reflect attack severity. Levels of sTNF-R75 were highly significantly associated with parasite density, indicating that this response is malaria-specific. The present study indicates that sTNF-R75 levels could become a useful immunological tool in malaria intervention studies, as they reflect changes in malaria-specific immune responses. Future studies should validate this potential in different endemic settings.
Mycobacteria-induced autoantibody production is associated with susceptibility to infection but not with host propensity to develop autoimmune disease
Authors: BRÁS A , ÁGUAS A P
Published: July 2010

Mycobacteria cause increase in autoantibody production in the host during the first weeks of infection. The level of the autoantibody enhancement varies widely in different hosts, suggesting that it depends on features of the host make-up. We have investigated the participation of two characteristics of the host in the modulation of mycobacteria-induced autoantibody production: (i) the host being genetically determined to later develop spontaneous autoimmune disease; (ii) the host being susceptible/resistant to mycobacterial infection. Mycobacterium avium infection was studied in 3-month-old mice that are prone (NZB and C57B1/6-lpr/lpr strains) or not (NZW and C.D2 strains) to develop, when older, autoimmune disease; these murine strains are either naturally susceptible (C57B1/6-lpr/lpr and NZW) or resistant (NZB and C.D2) to mycobacteria. Mycobacterium avium infection was produced by i.p. injection of 3 ± 107 viable bacilli. At days 15 and 30 of the infection, we determined the following parameters; (i) number of cells producing natural autoantibodies (splenic cells showing surface antibodies against bromelain-treated mouse (BrM) erythrocytes); (ii) suppression of the primary response to T cell-dependent antigen (i.e. to sheep erythrocytes); (iii) immunoglobulin classes and IgG isotypes; (iv) titres of anti-dsDNA antibodies; and (v) serum concentrations of interferon-gamma (IFN-γ). We found that the highest elevations in natural autoantibodies were associated with hosts being naturally susceptible to mycobacteria, but not with the host being genetically determined to later develop autoimmune disease. The rise in autoantibodies was predominantly of the IgM type, being associated with suppression of the T cell response and accompanied by increase in serum IFN-γ. Mycobacteria failed to induce any significant enhancement in pathogenic anti-dsDNA antibodies. Our data suggest that the finding of a high level of autoantibodies during the early phase of mycobacterial infection reflects host susceptibility to the infectious agent, and that it is not related with its propensity to later develop autoimmune disorders.
Human neutrophil Fc receptor-mediated adhesion under flow: a hollow fibre model of intravascular arrest
Authors: D'ARRIGO C , CANDAL-COUTO J J , GREER M , VEALE D J , WOOF J M
Published: July 2010

Human polymorphonuclear cells (PMN) were found to adhere to a novel model of blood vessel wall-associated IgG. The internal surfaces of cellulose acetate hollow fibres, of comparable internal diameter to small blood vessels, were coated with normal serum human IgG, heat-aggregated IgG (HAIgG), laminin or fibrinogen. Under conditions of flow mimicking those in a small vessel. PMN were found to adhere markedly only to immunoglobulin-coated fibres. Arrest on HAIgG was inhibited by excess soluble IgG but not by bovine serum albumin (BSA), demonstrating that the adhesion was IgG-specific and presumably mediated by FcγR on the PMN surface. Pre-adsorption of serum components onto HAIgG-coated fibres enhanced PMN arrest, due most probably to fixation of complement components by immobilized HAIgG, resulting in additional potential to entrap PMN via complement receptors such as CR3. Treatment of PMN with the regulatory neuropeptide substance P also enhanced adhesion to HAIgG-coated fibres and caused increased surface expression of FcγRI, FcγRII and FcγRIII. A mouse cell line derived from L cells, hR4C6, stably transfected with human FcγRII, was found to adhere under flow lo HAIgG-coated fibres, whilst untransfected parent L cells did not. This adhesion was similarly inhibited by excess soluble IgG, confirming the capability of FcγR to mediate cell arrest. The study strongly suggests that FcγR may play an important role in intravascular PMN arrest and we speculate that in inflammatory diseases PMN may adhere via FcγR to immobilized immunoglobulin on the vascular endothelium, with subsequent degranulation and tissue damage.
Spontaneous production of various cytokines except IL-4 from CD4+ T cells in the affected organs of sarcoidosis patients
Authors: Hoshino T , Itoh K , Gouhara R , Yamada A , Tanaka Y , Ichikawa Y , Azuma M , Mochizuki M , Oizumi K
Published: July 2010

We investigated surface antigens and spontaneous cytokine production of T cells from broncho-alveolar lavage fluid (BALF) and aqueous humor (AH) from pulmonary sarcoidosis patients for a better understanding of the role of T cells in granuloma formation. The levels of CD3, CD11b, and CD28 antigen expression on freshly isolated T cells in the BALF of patients were significantly lower than those in peripheral blood lymphocytes (PBL) of either sarcoidosis patients or healthy donors (HD). In contrast, the levels of CD80 (B7/B7-1) and CD86 (B70/B7-2) antigen expression were significantly higher on these T cells and alveolar macrophages in the BALF of patients. Fifty-three T cell clones (TCC) established from the BALF and AH of the three sarcoidosis patients displayed primarily either CD4+ CD11b+ CD28+ or CD4+ CD11b- CD28- phenotypes. Most (61–90%) of these TCC spontaneously produced greater amounts of IL-1a, IL-10, tumour necrosis factor (TNF), and granulocyte-macrophage colony-stimulating factor (GM-CSF) than did TCC from the PBL from sarcoidosis patients or HD (P < 0·05). Interferon-gamma (IFN-γ), IL-6, and IL-2, but not IL-4, were also produced by 40–48% of these TCC. These results suggest that CD4+ T cells of the affected organs of sarcoidosis patients are activated and involved in the immunopathogenesis of sarcoidosis through production of various cytokines.
Tolerance exists towards resident intestinal flora but is broken in active inflammatory bowel disease (IBD)
Authors: Duchmann R , Kaiser I , Hermann E , Mayet W , Ewe K , BÜSchenfelde K-H MEYER ZUM
Published: July 2010

Hyporesponsiveness to a universe of bacterial and dietary antigens from the gut lumen is a hallmark of the intestinal immune system. Since hyperresponsiveness against these antigens might be associated with inflammation, we studied the immune response to the indigenous intestinal microflora in peripheral blood, inflamed and non-inflamed human intestine. Lamina propria monocuclear cells (LPMC) isolated from inflamed intestine but not peripheral blood mononuclear cells (PBMC) of IBD patients with active inflammatory disease strongly proliferated after co-culture with sonicates of bacteria from autologous intestine (BsA), Proliferation was inhibitable by anti-MHC class II MoAb, suggesting that it was driven by antigen, LPMC from adjacent non-inflamed intestinal areas of the same IBD patients and PBMC or LPMC isolated from non-inflamed intestine of controls and patients with IBD in remission, in contrast, did not proliferate, PBMC or LPMC which had been tolerant to bacteria from autologous intestine, however, strongly proliferated after co-culture with bacterial sonicates from heterologous intestine (BsH). This proliferation was associated with an expansion of CD8+ T cells, increased expression of activation markers on both CD4+ and CD8+ lymphocyte subsets, and production of IL-12, interferon-gamma (IFN-γ), and IL-10 protein. These results show that tolerance selectively exists to intestinal flora from autologous but not heterologous intestine, and that tolerance is broken in intestinal inflammation. This may be an important mechanism for the perpetuation of chronic IBD.
Functional activity of the membrane-asssociated complement inhibitor CD59 in a pig-to-humanin vitromodel for hyperacute xenograft rejection
Authors: Heckl-ÖStreicher B , Binder R , Kirschfink M
Published: July 2010

Hyperacute rejection triggered by activation of the recipient's complement system represents the major barrier to successful xenotransplantation. Transfer of human membrane-associated complement regulators to donor organs has been suggested as one strategy to interfere with complement-mediated hyperacute xenograft rejection. Pigs are discussed as potential organ donors. We therefore investigated a putative protective function of the membrane-bound complement inhibitor CD59 in a pig-to-human in vitro model of hyperacute xenograft rejection. Aortic porcine endothelial cells were transfected with human CD59 cDNA. Expression of human CD59 was demonstrated by cytofluorimetric and RNA analysis. Removal of CD59 from the cell surface by phosphatidylinositol-specific phospholipase C (PI-PLC) demonstrated its production as a glycosyl phosphatidylinositol (GPI)-anchored protein. Functional activity of the transfected CD59 was tested by a lactate dehydrogenase (LDH) release assay for complement-mediated lysis. Porcine endothelial ceils expressing human CD59 were significantly protected from lysis by human serum complement compared with CD59- cells. The protective effect was abolished by preincubating the cells with anti-CD59 antibodies or PI-PLC. We calculated by Scatchard analysis that the established CD59+ cell line expressed a CD59 level comparable to that of human endothelial cells. Our results recommend the production of pigs transgenic for CD59.
Non-lymphoid and lymphoid cells in acute, chronic and relapsing experimental colitis
Authors: Palmen M J H J , Dieleman L A , Van Der Ende M B , Uyterlinde A , PeÑA A S , Meuwissen S G M , Van Rees E P
Published: July 2010

In rodents, intracolonic administration of ethanol 30% induces an acute colitis, while administration of 2,4,6-trinitrobenzene sulphonic acid (TNBS) in ethanol induces a longer lasting colitis. In the acute and chronic stages of experimental colitis, lymphoid and non-lymphoid cells were studied in the colon by immunohistochemistry. During the acute inflammation a high damage score of the colon was observed, which was related to an increase in the number of macrophages and granulocytes. Also a change in distributional patterns of macrophage subpopulations was found. The chronic stage of TNBS–ethanol-induced colitis was characterized by an increase in the number of lymphocytes, especially T cells. These data suggest that macrophages and granulocytes are important in the acute phase of experimental colitis, while lymphocytes play a pivotal role in the chronic stage. As most inflammatory bowel disease (IBD) patients have relapses during the chronic disease, we attempted to induce a relapse during experimental colitis by giving a second i.p. or s.c. dose of TNBS. This resulted in increased damage scores of the colon, new areas of ulceration and a further increase in macrophage numbers. No effect on the number of granulocytes was seen. These results indicate that it is possible to mimic relapses in experimental colitis by a second administration of TNBS, and suggest that the rats had been sensitized by the first dose of TNBS, given into the colon.
Autoimmunity in ulcerative colitis: tropomyosin is not the major antigenic determinant of the Das monoclonal antibody, 7E12H12
Authors: Hamilton M I , Bradley N J , Srai S K S , Thrasivoulou C , Pounder R E , Wakefield A J
Published: July 2010

Ulcerative colitis (UC) has a proposed autoimmune pathogenesis. A 40-kD antigen (P40) has been isolated from UC colon, bound to immunoglobulin. Tropomyosin has been reported as the target antigen of a MoAb (7E12H12) raised against P40. We set out to investigate whether tropomyosin is the major antigenic determinant for 7E12H12. Formalin-fixed, paraffin-processed and cryostat sections of fresh frozen colon from patients with UC, Crohn's disease and normals, were immunostained with 7E12H12 and commercial anti-tropomyosin antibodies. In addition, the immunoreactivity of 7E12H12 with cytoskeletal components was examined on human endothelial cells (HUVEC) using anti-tropomyosin as a positive control. Con-focal microscopy was used to determine the subcellular localization of signal. An extract of total colonic protein from UC colon was prepared. Using a combination of Western and immunoblotting (dot-blots), the immunoreactivities of both tropomyosin (porcine and chicken) and colon protein extract with either 7E12H12 or commercial anti-tropomyosin were examined. Immunocytochemically, 7E12H12 localized to the apical and basolateral regions of plasma membrane, and to the supranuclear cytoplasm in colonic epithelium. Using anti-tropomyosin antibody it was not possible to identify the cytoskeleton in colonic epithelium. Cytoskeletal components were identifiable in HUVEC cultures with anti-tropomyosin antibody but not with 7EI2H12. P40 antigen was identified in the colon protein extract by immunoblotting with 7E12H12. There was clear immunoreactivity between anti-tropomyosin antibody and both chicken and porcine tropomyosin, and the colon protein extract. 7E12H12 did not bind to either chicken or porcine tropomyosin in appropriately controlled systems. We conclude that the pattern of immunostaining with 7E12H12 is not cytoskeletal, and there is no reactivity in immunoblots, between tropomyosin and 7E12H12. Tropomyosin is not the major target antigen of this antibody in ulcerative colitis.
Chloroquine inhibits T cell proliferation by interfering with IL-2 production and responsiveness
Authors: Landewe R B M , Miltenburg A M M , Verdonk M J A , Verweij C L , Breedveld F C , Daha M R , Dijkmans B A C
Published: July 2010

Chloroquine (Chi) is an anti-rheumatic drug that is widely used in the treatment of rheumatoid arthritis (RA). It seems that T ceils are important in the pathogenesis of RA, but it is not known whether Chl acts via inhibition of T cell function. We here present evidence that Chl, just like cyclosporine A (CsA), inhibits Tcell proliferation as induced with immobilized αCD3 MoAb in a concentration-dependent manner, at least partly through interfering with the production of IL-2 protein and the induction of IL-2 inRNA. Furthermore, Chi impedes the responsiveness of T cell clones to IL-2 since (1) the inhibition of αCD3 MoAb-induced proliferation by Chi could not be reversed by rlL-2 and (2) Chi directly blocks IL-2-driven proliferation of cloned T cells. Chi appeared to interfere with the internalization (50% inhibition) and degradation (total blockade) of rIL-2. Finally, the combination of Chi and CsA synergistically inhibited T ceil proliferation. We conclude that Chi may inhibit functional properties of human T cells. although the drug is 100-to 1000-folds less potent than CsA in inhibiting T cell proliferation and IL-2 production, respectively. It is speculated that the in vitro. effects of Chl might be relevant in explaining the anti-rheumatic effect of this drug in patients with RA.
Immune reconstitution in human immunodeficiency virus type 1-infected children with different virological responses to anti-retroviral therapy
Authors: Anselmi A , Vendrame D , Rampon O , Giaquinto C , Zanchetta M , De Rossi A
Published: July 2010

Immune repopulation, despite virological failure, often occurs in children under highly active anti-retroviral therapy (HAART). The aim of this study was to analyse the characteristics of immune repopulation and activation in children with and without virological response to HAART. Fourteen human immunodeficiency virus type 1 (HIV-1)-infected children with suppression of HIV-1 plasma viraemia (virological responders, VR) and 16 virological non-responders (VNR) to therapy were studied at baseline and after approximately 2 years of HAART. During therapy, CD4+ T cells increased in both groups, but were higher in the VR than in the VNR group. All CD4+ T cell subsets (naive, central memory, effector/memory and CD38+) increased significantly in VR children, while there was a significant increase only in naive cells in VNR children. Naive CD8+ T cells and T cell receptor rearrangement excision circles (TREC), an indicator of thymic output, increased in both VR and VNR children. Activated CD8+CD38+ T cells decreased in VR but remained high in VNR children. Levels of circulating lipopolysaccharide (LPS), an indicator of microbial translocation, further increased in VNR children. In conclusion, HAART induced an increase in naive cells in all children, regardless of their virological response. However, the persistence of viraemia resulted in an impaired expansion of memory CD4+ T cells susceptible to HIV-1 infection, and together with the microbial translocation sustained the persistence of a high level of immune activation.
Programmed death-1 ligands-transfected dendritic cells loaded with glutamic acid decarboxylase 65 (GAD65) inhibit both the alloresponse and the GAD65-reactive lymphocyte response
Authors: He F-R , Zhu H-F , Huang H , Dai Y D , Shen X , Wang M , Li L , Xing W , Shen G-X
Published: July 2010

Type 1 diabetes (T1D) is due to a loss of immune tolerance to islet antigens, such as glutamic acid decarboxylase 65 (GAD65), for which islet transplantation is a promising therapy. Therefore, the generation of tolerance aiming at both alloantigen and GAD65 will help therapeutic intervention greatly in T1D. In this study, we tested the effect of programmed death-1 ligands (PD-L1)-transfected dendritic cells (DC) loaded with GAD65 on the alloresponse and GAD65-reactive lymphocyte response. The DC2·4 cell line was transfected with PD-L1 and co-cultured with GAD65. BALB-c mice were primed, respectively, by intraperitoneal injection with GAD65, PD-L1-transfected- or non-transfected DC (PD-L1/DC or DC), and PD-L1-transfected- or non-transfected DC loaded with GAD65 (PD-L1/DC/GAD65 or DC/GAD65). Splenocytes of treated mice were isolated and restimulated in vitro with GAD65 or the various DC populations above being used as stimulators, respectively. In the mixed lymphocyte reaction, DC/GAD65 were able to stimulate both allogeneic and GAD65-reactive lymphocytes. However, PD-L1/DC/GAD65 were poorer than DC/GAD65 at activating the GAD65-reactive lymphocyte response. Further, although PD-L1/DC could inhibit the alloresponse, PD-L1/DC/GAD65 were more effective at down-regulating the GAD65-reactive lymphocyte response. More importantly, PD-L1/DC/GAD65-primed lymphocytes exhibited the weakest proliferation when again restimulated in vitro by PD-L1/DC/GAD65. Additionally, PD-L1/DC/GAD65 down-regulated interferon-γ and up-regulated interleukin-10 production by activated lymphocytes. Therefore, combined stimulation in vivo and in vitro by PD-L1/DC/GAD65 could inhibit both the alloresponse and the GAD65-reactive lymphocyte response, which may contribute to controlling diabetes and islet transplant rejection.
The use of reflective and reasoned portfolios by doctors
Authors: Saltman Deborah C. , Tavabie Abdollah , Kidd Michael R.
Published: July 2010

Rationale Reflective portfolios have been widely trailed in vocational programmes for health sciences education. While not raised explicitly yet, there is recognition that an essential part of a portfolio may still be missing – that is the experience of theoretical and analytic questioning that should occur after reflection.

Aims and objectives In this paper, we argue that portfolios require both reflective and reasoned components to provide a more complete range of educational experiences for postgraduate doctors in the pursuit of higher qualifications.

Methods We describe the differentiating features between the two components and conclude with some suggestions on how both reflective and reasoned components of portfolios may work synergistically through a series of case studies.

Results The reflective style of portfolio, which is in current use, has been widely accepted as a major advance in postgraduate medical education at all levels. It provides an effective vehicle for encouraging self‐consideration and generalizability. It is an important adjunct where it functions as a personal development tool as well as a sophisticated log book. A reasoned or pedagogic element encourages more analytic activities: adding an additional component that can ground the portfolio in theory or research. The portfolio is then able to function in a range of academic spheres and provide a theoretical platform for the wide range of non‐clinical empirical work undertaken in clinical settings. The addition of a reasoned component also increases the emphasis on the introduction of new concepts outside the sphere of the learner's experience.

Conclusions A portfolio containing both reflective and reasoned components is then able to function in a range of academic spheres and provide a theoretical platform for the wide range of non‐clinical empirical work undertaken in clinical settings. In an iterative mode, such a portfolio element can support a forum for questioning established concepts within a pedagogical environment – the essence of evidence‐based practice.
Studies on Recombination within the Mouse H‐2 Gene Complex. III. Further Serological Analyses of the H‐2t Haplotypes*
Authors: David Chella S. , Hansen Ted H. , Shreffler Donald C.
Published: July 2010

The five H‐2t haplotypes are all intra‐H‐2 recombinants which have identical K and D regions, but differ in the central regions of the complex. Analyses of H‐2, Ia and Ss‐Slp types indicate that the crossover in H‐2t1 took place between the K and I regions, in the H‐2t2 between G and D, in H‐2t3 and H‐2t4 between the Z‐B and I‐C subregions and in H‐2t5 between I and S. The set of recombinants will be very valuable for studies of traits which map in the I and S regions.
Histocompatibility Antigens and Myocardial Infarction
Authors: Scott Brian B. , McGuffin P. , Rajah S. M. , Stoker J. B. , Losowsky M. S.
Published: July 2010

The incidence of histocompatibility (HLA) antigens in 100 men with myocardial infarction was compared with the incidence in 500 healthy controls. No significant difference was found, providing no support for either a genetic or an immunological basis for myocardial infarction.