bluuebunny/crossref_metadata_2025 huggingface dataset:
Immunological study on CD3 defective cutaneous T cell lymphoma cells from a patient with Sézary syndrome
Authors: Sano S , Matsui Y , Itami S , Yoshikawa K
Published: March 2003

Here we investigated the nature of cutaneous T cell lymphoma (CTCL) cells lacking surface CD3. A large number of CD3−CD4 T cells were found in the peripheral blood and lesional skin of a patient with Sézary syndrome, which is a variant of CTCL. Southern blot analysis revealed that a clonal rearrangement of T cell receptor (TCR) genes was detected in the separated CD3−CD4 cells, whereas CD3+CD4 cells showed no clonal rearrangement, indicating that the CD3−CD4 cells represented CTCL cells. However, the CTCL cells expressed TCR with a particular Vβ apart from CD3. The CTCL cells showed significant responses to staphylococcal enterotoxins (SEs) in vitro, although they hardly responded to phytohaemagglutinin, Mycobacterium tuberculosis antigen, and alloantigen. They required antigen-presenting cells (APC) to respond to SEB. Blocking analyses with MoAbs revealed that they recognized SEB through TCR depending on HLA-DR and intercellular adhesion molecule-1 (ICAM-1). Taken collectively, these results indicate that the CTCL cells lacking surface CD3 could proliferate in response to bacterial superantigens, whereas the responses to conventional antigens were generally suppressed. These results also implied that CTCL could be exacerbated by bacterial infection.
A dominant role for non-MHC gene effects in susceptibility to cyclosporin A (CsA)-induced autoimmunity
Authors: Damoiseaux J G M C , Beijleveld L J J , Van Breda Vriesman P J C
Published: March 2003

Lethally irradiated LEW rats reconstituted with syngeneic bone marrow and given CsA for a 4-week period develop a graft-versus-host-like disease upon withdrawal of CsA. This T cell-mediated autoimmune disease is referred to as CsA-induced autoimmunity (CsA-AI). CsA-AI-susceptible LEW rats and resistant BN rats differ greatly in the composition of their peripheral T cell compartment. To dissect the role of MHC and non-MHC genes in the development of peripheral T cell subsets in combination with susceptibility to CsA-AI the respective MHC congenic strains (LEW-1N and BN-1L) were examined for their T cell subsets and for their ability to develop CsA-AI. In this study we show that the Th1/Th2-like cell ratio as well as susceptibility to CsA-AI are under control of the non-MHC genes. This suggests that the Th1/Th2-like cell ratio is a critical determinant for development of CsA-AI. Alternatively, resistance can be attributed to lack of target organ susceptibility due to the absence of the target autoantigen in resistant rat strains. This interpretation is rejected, since both BN as well as BN-1L rats consistently develop the characteristic macroscopic and microscopic signs of CsA-AI upon adoptive transfer with autoreactive LEW-1N and LEW T cells, respectively. Therefore, it can be concluded that the non-MHC genes encode for immune deviation and thereby determine susceptibility or resistance to CsA-AI.
Chemokine profiles in the cerebrospinal fluid (CSF) during the course of pyogenic and tuberculous meningitis
Authors: Mastroianni C M , Lancella L , Mengoni F , Lichtner M , Santopadre P , D'agostino C , Ticca F , Vullo V
Published: March 2003

The concentrations of the chemokines IL-8, monocyte chemotactic protein-1 (MCP-1) and macrophage inflammatory protein-1α (MIP-1α) were measured in 120 CSF samples from 23 patients with pyogenic meningitis and from 11 patients with tuberculous meningitis (TBM) and in 10 CSF from subjects with non-infectious neurological diseases. The chemokine concentrations in patients with meningitis were significantly higher than in control subjects (P < 0.0001). The highest CSF levels were found for IL-8 (median 2917 pg/ml) and MCP-1 (median 2557 pg/ml), whereas those of MIP-1α were less significantly elevated (median 24 pg/ml) (P < 0.0001). Patients with pyogenic meningitis had higher levels of IL-8 and MCP-1 than those with TBM (P < 0.0001). In serial samples from patients with pyogenic meningitis IL-8 levels declined before MCP-1 and MIP-α. In the case of TBM, IL-8, MCP-1 and MIP-1α decreased more gradually during treatment and were detectable in the CSF for several weeks, without any characteristic time course of elimination. These data indicate that patients with pyogenic meningitis and TBM show different chemokine profiles in CSF. The distinct chemokine pattern could be responsible for a differential attraction and activation of leucocytes in the CSF which is reflected in differences in the inflammatory response and clinical course of pyogenic meningitis and TBM.
Apoptosis of Th1-like cells in experimental tuberculosis (TB)
Authors: DAS G , VOHRA H , SAHA B , AGREWALA J N , MISHRA G C
Published: March 2003

Th1 cell-induced anti-mycobacterial immunity is lost during a progressive Mycobacterium tuberculosis infection in a susceptible host. This study was designed to test the mechanism of the loss of anti-mycobacterial cell-mediated immune response. We demonstrate that M. tuberculosis infection results in increased Fas expression and decreased Bcl-2 expression in CD4+ T cells. When CD4+ T cells are stimulated in vitro, they show increased apoptosis and decreased production of IL-2 and interferon-gamma (IFN-γ) but not of IL-4. These changes may result in selective apoptosis of Th1-like cells, leading to the loss of cell-mediated immune response against M. tuberculosis.
Serum concentrations of cytokines in patients with active tuberculosis (TB) and after treatment
Authors: VERBON A , JUFFERMANS N , VAN DEVENTER S J H , SPEELMAN P , VAN DEUTEKOM H , VAN DER POLL T
Published: March 2003

During TB cytokines play a role in host defence. To determine the cytokine pattern during various disease stages of TB, serum levels of IL-12, interferon-gamma (IFN-γ), IL-4, IL-6 and IL-10 were measured in 81 patients with active TB, 15 patients during therapy and 26 patients after anti-tuberculous therapy as well as in 16 persons who had been in close contact with smear-positive TB and in 17 healthy controls. IFN-γ was elevated during active TB when compared with healthy controls, declining during and after treatment. IL-12 (p40 and p70) serum levels were not significantly higher in patients with active TB compared with any of the other groups. IL-4 levels were low in all groups. IL-6 and IL-10 serum levels were elevated in patients with active TB and during treatment. In patients with active TB serum levels of IFN-γ and IL-6 were higher in patients with fever, anorexia and malaise. IL-12 levels were higher in patients with a positive smear. Cytokine levels did not correlate with localization of TB (pulmonary versus extrapulmonary), or skin test positivity. Cytokines directing a Th1 response (IL-12) or a Th2 response (IL-4) were not elevated in sera of this large group of patients with pulmonary and extrapulmonary TB. In patients with active TB, cytokines that were elevated in serum were IFN-γ, IL-6 and IL-10.
Regulation of CR3 (CD11b/CD18)-dependent natural killer (NK) cell cytotoxicity by tumour target cell MHC class I molecules
Authors: VĚTVIČKA V , HANIKÝŘOVÁ M , VĚTVIČKOVÁ J , ROSS G D
Published: March 2003

Phagocyte and NK cell CR3 functions as both an adhesion molecule and an iC3b receptor mediating cytotoxic responses to microorganisms. Cytotoxic activation of iC3b receptor function requires ligation of both a CD11b I-domain site for iC3b and a lectin site located in the C-terminus of CD11b. Because tumours lack the CR3-binding polysaccharides of bacteria and fungi, iC3b-opsonized tumours do not stimulate CR3-dependent cytotoxicity. Previous studies showed that NK cells could be induced to kill iC3b-opsonized tumours with small soluble β-glucans that bound with high affinity to CR3, bypassing the absence of similar polysaccharides on tumour membranes. Because CR3 signalling requires several tyrosine phosphorylation events, it appeared possible that CR3-dependent killing of autologous tumour cells might be suppressed by NK cell inhibitory receptors for MHC class I (KIR and CD94/NKG2) whose action involves recruitment of SHP-1 and SHP-2 tyrosine phosphatases. In the current study, Epstein–Barr virus (EBV)-transformed B cells were used as targets following opsonization with iC3b. Soluble β-glucan primed CR3 for killing of iC3b-coated B cells, but autologous class I-bearing targets were 84% more resistant than class I-deficient Daudi cells. Blockade of target cell class I with a MoAb specific for a domain recognized by both KIR and CD94/NKG2 resulted in comparable killing of class I+ B cells. By contrast, another MoAb to class II had no effect on cytotoxicity. These data suggest that NK cell recognition of class I suppresses CR3/tyrosine kinase-dependent cytotoxicity in the same way as it suppresses cytotoxicity mediated by other tyrosine kinase-linked receptors such as FcγRIIIA (CD16).
Expression of killer inhibitory receptors on cytotoxic cells from HIV-1-infected individuals
Authors: GALIANI M D , AGUADO E , TARAZONA R , ROMERO P , MOLINA I , SANTAMARIA M , SOLANA R , PEÑA J
Published: March 2003

Dysfunction of cytotoxic activity of T and natural killer (NK) lymphocytes is a main immunological feature in patients with AIDS, but its basis are not well understood. It has been recently described that T and NK cell-mediated cytotoxicity can be regulated by HLA killer inhibitory receptors (KIR). In this work, we have determined on cytotoxic T cells and NK cells from HIV-1-infected individuals the expression of the following KIR molecules: p58, p70, and ILT2 (immunoglobulin-like family KIR) as well as CD94 and NKG2A (C-lectin-type family KIR). With some exceptions, no significant changes were found on the expression of immunoglobulin-like KIR in either CD8+ or CD56+ cells. Interestingly, the percentages of CD8+ and CD56+ cells expressing CD94 were significantly increased in these individuals. We also show that, in vitro, IL-10 up-regulates CD94 expression on CD8+ and CD56+ cells obtained from normal individuals, suggesting that the augmented expression observed in HIV-infected individuals could be related to the high levels of IL-10 previously described in HIV-1-infected individuals.
Evidence for control of tumour necrosis factor-alpha (TNF-α) activity by TNF receptors in patients with proliferative diabetic retinopathy
Authors: LIMB G A , SOOMRO H , JANIKOUN S , HOLLIFIELD R D , SHILLING J
Published: March 2003

TNF-α has been implicated in the pathogenesis of insulin- dependent diabetes mellitus (IDDM). At present there are no studies linking serum levels of soluble TNF receptors (sTNF-R) to the development of diabetic microvascular complications such as proliferative diabetic retinopathy (PDR), or to the production of TNF-α in these patients. We investigated serum levels of sTNF receptors (sTNF-RI and sTNF-RII) in IDDM patients with or without PDR, and related these to the in vitro production of TNF-α upon activation of whole blood and isolated mononuclear cells (MNC). We observed higher serum levels of sTNF-RI in IDDM patients with active (range 945–6630 pg/ml; P = 0.029) or quiescent PDR (range 1675–4970 pg/ml; P = 0.00092) than in individuals with IDDM without retinopathy (range 657–2617 pg/ml) or healthy controls (range 710–1819 pg/ml; P = 0.0092 and 0.0023, respectively). Increased serum levels of sTNF-RII were also seen in IDDM patients with active PDR (range 1749–5218 pg/ml; P = 0.034) or quiescent PDR (range 1494–5249 pg/ml; P = 0.0084) when compared with disease controls (range 1259–4210 pg/ml) or healthy subjects (range 1237–4283 pg/ml). Whole blood production of biologically active TNF-α was lower in PDR patients than in disease (P = 0.04) and healthy controls (P < 0.005), contrasting with a higher production of TNF-α by lipopolysaccharide (LPS)-activated MNC from PDR patients (P = 0.013). Inhibition of TNF-α by TNF-R in plasma supernatants of activated blood from PDR patients was demonstrated by increase of TNF-α activity in the presence of anti-TNF-RI and anti-TNF-RII antibodies. These observations suggest that abnormalities in TNF-α production and control may operate during the development of microvascular complications of diabetes mellitus.
Increased interferon-gamma (IFN-γ) levels producedin vitroby alloactivated T lymphocytes in systemic sclerosis and Raynaud's phenomenon
Authors: MOLTENI M , DELLA BELLA S , MASCAGNI B , BAZZI S , ZULIAN C , COMPASSO S , LESSI M , SCORZA R
Published: March 2003

The aim of the present study was to analyse the in vitro proliferation and cytokine production by alloantigen-stimulated peripheral blood mononuclear cells (PBMC) obtained from patients affected by systemic sclerosis (SSc) and patients with Raynaud's phenomenon (RP). In SSc patients the proliferation of PBMC stimulated in vitro with alloantigens was significantly increased compared with healthy subjects, while no differences were observed for RP patients. Lymphocytes from SSc patients also produced larger amounts of IFN-γ compared with healthy controls. However, patients with clinically active disease had lower IFN-γ levels than those found in clinically stable patients. Patients affected by RP showed significantly higher levels of IFN-γ than healthy subjects. Analysis at the clonal level of the lymphocyte subsets involved in alloantigen stimulation in one patient affected by active SSc, and one subject with RP confirmed the results obtained using PBMC. In particular, in the RP patient but not in the SSc patient, we observed a population of CD4+ T cells which proliferated to alloantigens in vitro and produced high levels of IFN-γ. We suggest that T lymphocytes producing high levels of IFN-γ might play a protective role in RP patients and in established scleroderma.
Detection of antibodies directed against the cytoplasmic region of the human acetylcholine receptor in sera from myasthenia gravis patients
Authors: TZARTOS S J , REMOUNDOS M
Published: March 2003

The nicotinic acetylcholine receptor (AChR) is the autoantigen in the human autoimmune disease myasthenia gravis (MG). Anti-AChR antibodies in MG sera bind mainly to conformational epitopes, therefore the determination of their specificities requires the use of native AChR. Antibody competition studies suggest that most MG antibodies are directed against the extracellular part of the molecule, whereas antibodies directed against the cytoplasmic region of the AChR have not been detected. To determine whether even small quantities of such antibodies exist in MG sera, we performed competition experiments based on the inhibition by MG sera of the binding of MoAbs to the human AChR, rather than inhibition by MoAbs of the binding of MG sera performed earlier. When MoAbs directed against cytoplasmic epitopes on the α or β subunits (α373–380 and β354–360) were used as test MoAbs, 17% or 9% of MG sera inhibited the binding of the anti-α or anti-β subunit MoAbs, respectively, by ≥ 50%. Non-specific inhibition was excluded. These results suggest the presence, in several MG sera, of antibodies directed against cytoplasmic regions of the AChR; yet these antibodies seemed to represent a relatively small proportion of the total anti-AChR antibodies. The corresponding epitopes may be involved in the inducing mechanisms in certain MG cases, and knowledge of the presence of such antibodies may be useful in understanding the autoimmune mechanism involved in MG.
The effects of extracorporeal whole body hyperthermia on the functional and phenotypic features of canine peripheral blood mononuclear cells (PBMC)
Authors: KEARNS R J , RINGLER S , KRAKOWKA S , TALLMAN R , SITES J , OGLESBEE M J
Published: March 2003

In this study the effect of transient 42.3°C whole body hyperthermia (WBH) on the distribution of PBMC phenotypes and in vitro blastogenic responsiveness was determined in dogs. Hyperthermia (n = 6) was induced by heating venous blood during extracorporeal circulation (venous perfusion WBH); perfused non-heated dogs (n = 4) were used as controls. Both euthermic and hyperthermic perfusion produced transient lymphopenia which normalized in controls after perfusion but persisted in hyperthermic animals throughout the 8-day post-perfusion observation interval. The transient lymphopenia in control dogs was non-selective. In contrast, WBH-associated lymphopenia was selective, in that CD5+ T lymphocytes were more sensitive to hyperthermia than sIg+ B cells and, within the T cell compartment, suppressor (CD8+) cells were more sensitive to hyperthermic stress than helper (CD4+) lymphocytes. Functional analyses showed that WBH caused persistent suppression of PBMC blastogenesis in response to T cell phytomitogens. Increased plasma cortisol levels were correlated to peak lymphopenia and hyporesponsiveness to phytomitogens. Despite these alterations, high grade WBH was well tolerated and there was no evidence of opportunistic infection.
Gene expression of 5-lipoxygenase and LTA4 hydrolase in renal tissue of nephrotic syndrome patients
Authors: MENEGATTI E , ROCCATELLO D , FADDEN K , PICCOLI G , DE ROSA G , SENA M , RIFAI A
Published: March 2003

Leukotrienes (LT) of the 5-lipoxygenase pathway constitute a class of potent biological lipid mediators of inflammation implicated in the pathogenesis of different models of experimental glomerulonephritis. The key enzyme, 5-lipoxygenase (5-LO), catalyses oxygenation of arachidonic acid to generate the primary leukotriene LTA4. This LT, in turn, serves as a substrate for either LTA4 hydrolase, to form the potent chemoattractant LTB4, or LTC4 synthase, to produce the powerful vasoconstrictor LTC4. To investigate the potential role of LT in the pathogenesis of human glomerulonephritis with nephrotic syndrome, we examined the gene expression of 5-LO and LTA4 hydrolase in renal tissue of 21 adult patients with nephrotic syndrome and 11 controls. The patients consisted of 11 cases of membranous nephropathy (MN), seven focal and segmental glomerulosclerosis (FSGS), two non-IgA mesangial glomerulonephritis and one minimal change disease. Total RNA purified from renal tissue was reverse transcribed into cDNA and amplified with specific primers in a polymerase chain reaction (RT-PCR). Eight patients' renal tissue, four MN and four FSGS, co-expressed 5-LO and LTA4 hydrolase. In situ hybridization analysis revealed 5-LO expression and distribution limited to the interstitial cells surrounding the peritubular capillaries. Comparative clinical and immunohistological data showed that these eight patients had impaired renal function and interstitial changes that significantly correlated with 5-LO expression. These findings suggest that leukotrienes may play an important role in the pathogenesis of MN and FSGS. These results are also relevant to elucidating the pathophysiologic mechanisms which underlie progression to renal failure in these diseases.
Cytokine-producing cells in peripheral blood of children with coeliac disease secrete cytokines with a type 1 profile
Authors: HANSSON T , DANNÆUS A , KLARESKOG L
Published: March 2003

Coeliac disease (CoD) is a small intestinal disorder characterized by crypt cell hyperplasia and villous atrophy, and the production of cytokines from T cells and macrophages are of importance for the histological changes seen in CoD. A peroral immunization with an antigen, which gives rise to a mucosal immune response, may increase the levels of circulating cytokine-producing cells, and we wanted to obtain a better picture of an eventual emergence of activated circulating T cells in the peripheral blood in children with CoD. The cytokine expression of interferon-gamma (IFN-γ), IL-4, IL-6 and IL-10 was measured at the single-cell level by an ELISPOT method in 38 children with CoD. The numbers of IFN-γ-producing cells in the peripheral blood was increased in children with untreated CoD (P < 0.01) and after gluten challenge (P < 0.05) compared with healthy controls. Also, the numbers of IL-6-producing cells were increased (P < 0.05) after gluten challenge compared with the healthy controls. A paired comparison showed that the numbers of IFN-γ-producing cells increased after gluten challenge (P < 0.05), whereas no such change was seen for IL-4- or IL-10-producing cells. There were no differences in the numbers of IFN-γ-producing cells between the group of children with treated CoD and the groups of untreated or challenged CoD children. IL-4 production correlated with serum levels of total IgE. These results show that circulating mononuclear cells in children with active CoD secrete cytokines compatible with a type 1 response.
Absence of significant Th2 response in diabetes-prone non-obese diabetic (NOD) mice
Authors: HARTEMANN A H , RICHARD M-F , BOITARD C
Published: March 2003

Accumulating evidence suggests that Th1 T cells play a pivotal role in the development of autoimmune diabetes. Conversely, promoting a Th2 response inhibits disease progression. However, it has not been determined whether Th2 cells are regulatory T cells that fail at the time of diabetes development in naive non-diabetic NOD mice. Therefore, in order to evaluate cytokine secretion by spleen and islet infiltrating T cells in NOD mice at different stages of the autoimmune process, we developed an ELISPOT assay that detects IL-2, IL-4, and interferon-gamma (IFN-γ) secretion in vitro at the single-cell level. We showed that, whatever the age considered, IFN-γ is predominantly secreted, and that no IL-4-secreting cells are detected in the islets of male and female NOD mice. Spleen cells from 8-week-old female NOD mice, which include regulatory suppressor T cells, do not secrete IL-4, either upon presentation of islet cell antigens in vitro, or after transfer in vivo, but do secrete IFN-γ. IFN-γ secretion by T cells from diabetic mice results from CD4 but not CD8 T cells in transfer experiments into NOD/severe combined immunodeficient (SCID) recipients. These results suggest that (i) detection of regulatory CD4 T cells in NOD mice is not paralleled by a Th2 response; (ii) β cell destruction does not depend on a switch from a Th2 to a Th1-type response; and (iii) CD8 T cells do not participate in induction of diabetes by secreting IFN-γ.
Mode of delivery directs the phagocyte functions of infants for the first 6 months of life
Authors: GRÖNLUND M M , NUUTILA J , PELTO L , LILIUS E M , ISOLAURI E , SALMINEN S , KERO P , LEHTONEN O P
Published: March 2003

Factors that direct the immune responsiveness of the newborn beyond the immediate post-natal period are not known. We investigated the influence of mode of delivery and type of feeding on the phagocyte activity during the first 6 months of life. Sixty-four healthy infants (34 delivered vaginally and 30 by elective Caesarean section) were studied at birth and at the ages of 2 and 6 months. Phagocyte functions were studied by measuring the chemiluminescence (CL) activity of whole blood and isolated leucocytes and by investigating the expression of phagocyte receptors (FcγRI (CD64), FcγRII (CD32), FcγRIII (CD16), CR1 (CD35), CR3 (CD11b) and FcαR (CD89)) on neutrophils, monocytes and eosinophils by using receptor-specific MoAbs and immunofluorescence flow cytometry. Infants born by elective Caesarean section had significantly higher CL activity than those delivered vaginally during the entire 6-month follow up. In addition, infants who received formula feeds had significantly higher CL activity at 6 months of age and higher expression of FcγRI-, Fcα- and CR3-receptors on neutrophils than infants exclusively breast-fed. We suggest that stress reaction associated with labour influences the phagocytic activity measured in the cord blood but later during infancy the intraluminal antigens, gut microflora and diet, become important determinants in immune programming of human individuals.
Analysis of Epstein–Barr virus (EBV) receptor CD21 on peripheral B lymphocytes of long-term EBV− adults
Authors: JABS W J , PAULSEN M , WAGNER H J , KIRCHNER H , KLÜTER H
Published: March 2003

Primary infections with EBV are rarely observed after the age of 20. Some individuals even remain seronegative all their lives. Previously, a lack of EBV receptors on B cells of persistently EBV− adults was described as a reason for long-term EBV-seronegativity. The present study examined the CD21 receptor status of 20 repeatedly EBV− healthy adults and 32 EBV+ volunteers by means of flow cytometry. CD21 molecules on the surface of CD19+ B cells were quantified using anti-IgG-coated microbeads. The percentage of CD19+/CD21+ B lymphocytes was slightly lower in the peripheral blood of EBV−donors, but the CD21 antibody binding capacity on CD19+ B cells showed no significant differences between EBV− and EBV+ adults. In vitro studies showed an equally good EBV transformability of peripheral B lymphocytes of EBV− and EBV+ donors. Since HLA-DR was recently described as a co-receptor for EBV infection of B cells, we also determined HLA-DRB1 alleles in the EBV−group. We found a significant negative association of EBV-seronegativity with HLA-DR13 in comparison with 111 healthy blood donors. In summary, a biologically significant lack of the EBV receptor CD21 on peripheral B lymphocytes of persistently EBV− adults was excluded as a reason for long-term EBV-seronegativity.
Tumour necrosis factor-alpha (TNF-α) transcription and translation in the CD4+ T cell-transplanted scid mouse model of colitis
Authors: WILLIAMS A M , WHITING C V , BONHAGEN K , REIMANN J , BREGENHOLT S , CLAESSON M H , BLAND P W
Published: March 2003

The adoptive transfer of activated CD4+α/β T cell blasts from the spleens of immunocompetent C.B-17+/+ or BALB/cdm2 mice into C.B-17scid/scid (scid) mice induces a colitis in the scid recipient within 8 weeks, which progresses to severe disease within 16 weeks. T cells isolated from recipient colon show a Th1 cytokine phenotype. We have examined the relationship between the phenotype of the cellular infiltrate and the transcription and translation of the proinflammatory cytokine TNF-α. The techniques of double indirect immunohistology and in situ hybridization using digoxigenin-labelled riboprobes were used. The prominent myeloid cell infiltrate in diseased tissues comprised F4/80+, Mac-l+ macrophages, neutrophils, dendritic cells and activated macrophages. TNF-α transcription and translation were associated with activated macrophages in the lamina propria. Activated macrophages transcribing and translating TNF-α were clustered in areas of tissue destruction. Crypt epithelium of inflamed tissues transcribed TNF-α at a very early stage of the disease process, but translation of TNF-α protein could only be found in advanced epithelial dysplasia. This indicates differential post-transcriptional control of TNF-α in activated macrophages and the epithelium.
Rarity of autoantibodies to a major autoantigen, thyroid peroxidase, that interact with denatured antigen or with epitopes outside the immunodominant region
Authors: GUO J , WANG Y , JAUME J C , RAPOPORT B , MCLACHLAN S M
Published: March 2003

The nature of the autoantibody repertoire to the dominant autoantigen in human autoimmune thyroid disease is controversial. There is evidence that autoantibodies to thyroid peroxidase (TPO) interact with overlapping conformational epitopes in an immunodominant region and binding to denatured (DN) protein is decreased. Contrary data demonstrate TPO autoantibody reactivity with DN-TPO or polypeptide fragments. However, none of the TPO-specific, human monoclonal autoantibodies isolated to date preferentially recognize denatured autoantigen. We therefore searched an immunoglobulin gene phage display library for human autoantibodies that bind TPO denatured by reduction and alkylation (DN-TPO). Thyroid-infiltrating B cells from a typical TPO autoantibody-positive patient were the source of mRNA for library construction. Surprisingly, the library enriched after panning on DN-TPO, as well as a panel of individual clones, preferentially bound native (N)-TPO. Of 13 clones selected using DN-TPO or N-TPO, 12 clones recognized the TPO immunodominant region. Moreover, regardless of selection with N-TPO or DN-TPO, their heavy and light chains were encoded by similar VDJ and Vκ combinations. One clone (DN4), isolated using DN-TPO, did not interact with the TPO immunodominant region and its H chain derives from a different VH gene. Although DN4 binds specifically to TPO, its affinity is low, unlike the high affinities of other human TPO autoantibodies. In conclusion, human monoclonal autoantibodies that preferentially recognize denatured TPO could not be isolated from an immunoglobulin gene library despite selection with denatured protein. Our findings demonstrate the bias of the human B cell repertoire towards recognition of an immunodominant region on the conformationally intact form of a major thyroid autoantigen.
Enhancing of anti-viral activity against HIV-1 by stimulation of CD8+ T cells with thymic peptides
Authors: MÜLLER H , MAYER G , BEHNKE B , HEIMÜLLER E , HAMSCHER G , IMMLER D , SIETHOFF C , MEYER H E , SCHREIBER M
Published: March 2003

HIV-1 can be neutralized by soluble factors produced and secreted by activated CD8+ T cells. Production of such anti-viral CD8 factors (including chemokines) can be induced with IL-2 or phytohaemagglutinin (PHA). In addition to PHA or IL-2, we have co-stimulated CD8+ T cells with PHA/IL-2 and a mixture of thymic peptides (TP) of molecular weights below 10 kD. For the activation, CD8+ T cells were purified from peripheral blood mononuclear cells of HIV-1− individuals and any resultant anti-viral activity was monitored using an HIV-1 neutralization assay. Using HIV-1 isolates highly resistant to chemokine inhibition we detected significantly higher levels of HIV-1 neutralizing activity in CD8+ T cell culture supernatants which had been co-activated with TP. When the TP-induced anti-viral activity was monitored, neutralization of both non-syncytia-inducing (NSI) and syncytia-inducing (SI) patient isolates was enhanced by 38% (NSI, PHA +/− TP), 66% (SI, PHA +/− TP), 28% (NSI, IL-2 +/− TP), and 57% (SI, IL-2 +/− TP) compared with the anti-viral activity present in supernatants from CD8+ T cell cultures stimulated only with PHA or IL-2. Peptide sequence analysis of purified TP showed that the TP mixture predominantly contains peptides with homology to human histone and collagen sequences. Our data demonstrate that CD8+ T cells are additionally activated by a mixture of TP. In this way, the production of HIV-1 neutralizing CD8 factors can be enhanced.
Reduced IL-4 and interferon-gamma (IFN-γ) expression by CD4 T cells in patients with chronic lymphocytic leukaemia
Authors: HILL S J , PETERS S H , AYLIFFE M J , MERCEICA J , BANSAL A S
Published: March 2003

CD7 co-expression by CD4 T cells has been reported to be higher in the Th1 compared with the Th2 functional subset. Clinical immunodeficiency and immune dysregulation are more prevalent in the advanced stages of B cell chronic lymphocytic leukaemia (B-CLL). To analyse this further 25 patients with B-CLL and 11 healthy subjects were examined for cell surface CD7 and intracellular IFN-γ and IL-4 expression in the peripheral blood CD4+ T helper cell population. Significantly decreased CD7, IFN-γ and IL-4 expression was observed in the patients with B-CLL (P < 0.001). While CD7 negativity and IL-4 expression were more frequent in the later stages of the disease, this did not attain statistical significance. These results suggest a possible explanation for the reduced cellular and humoral immunity in B-CLL.
Cytokine release by ovine macrophages following infection withChlamydia psittaci
Authors: Entrican G , Wilkie R , Mcwaters P , Scheerlinck J P , Wood P R , Brown J
Published: March 2003

Chlamydia psittaci is an obligate intracellular pathogen that causes abortion in both sheep and humans. The disease in sheep (but not humans) is characterized by a long-term persistent phase that appears to be under the control of interferon-gamma. However, nothing is known about cytokine induction that precedes the persistent phase in sheep. Primary alveolar lavage cells recovered from normal adult sheep were used to study cytokine production in the first 72 h of infection with C. psittaci. These cells were phenotypically characteristic of macrophages, being adherent, phagocytic, CD14+ and staining positive for non-specific esterase. In vitro infection of the macrophages with C. psittaci resulted in the release of IL-1β, IL-8 and granulocyte-macrophage colony-stimulating factor (GM-CSF) as measured by ovine-specific ELISAs. Heat-treated chlamydiae (1 h at 65°C) did not induce the release of IL-1β, but the release of IL-8 was similar to that induced by untreated organisms. The cells from different sheep varied most notably in their patterns of GM-CSF release in response to heat-treated and untreated organisms.
Inhibition of dextran sulphate sodium (DSS)-induced colitis in mice by intracolonically administered antibodies against adhesion molecules (endothelial leucocyte adhesion molecule-1 (ELAM-1) or intercellular adhesion molecule-1 (ICAM-1))
Authors: Hamamoto N , Maemura K , Hirata I , Murano M , Sasaki S , Katsu K
Published: March 2003

We examined the effect of intracolonic administration of anti-adhesion molecule antibodies on DSS-induced colitis in mice. Immunohistochemical staining in mice with colitis showed increased expression of ELAM-1 and ICAM-1 on endothelial cells of vessels in the lamina propria and submucosa at sites of inflamed lesions. Intracolonic administration of anti-ELAM-1 or anti-ICAM-1 antibody decreased bloody stools, anaemia, and histologically evident damage, as well as myeloperoxidase activity and IL-1β content. We concluded that adhesion molecule expression is important in the development of DSS-induced colitis in mice and that intracolonic administration of anti-adhesion molecule antibodies, especially anti-ELAM-1 antibody, effectively inhibits the colonic inflammation. Intracolonic administration of anti-adhesion molecule antibodies may show therapeutic promise in ulcerative colitis.
Quantitative analysis of peripheral blood Th0, Th1, Th2 and the Th1:Th2 cell ratio during normal human pregnancy and preeclampsia
Authors: Saito S , Sakai M , Sasaki Y , Tanebe K , Tsuda H , Michimata T
Published: March 2003

We calculated the percentage of Th1, Th2, Th0 cells and the Th1:Th2 cell ratio of peripheral blood from normal pregnant subjects and preeclampsia patients using flow cytometry which can analyse both the surface marker, CD4, and intracellular cytokines, interleukin (IL)-4 and interferon (IFN)-γ. In normal pregnancy, the percentage of Th1 cells was significantly lower in the third trimester, and the ratios of Th1:Th2 were significantly lower in the second and third trimester than in nonpregnant subjects. In contrast, the percentage of Th1 cells and the ratios of Th1:Th2 in preeclampsia were significantly higher than in normal third trimester pregnant subjects. The percentage of Th2 cells in preeclampsia was significantly lower than in third trimester of normal pregnancy. Additionally, peripheral blood mononuclear cells from these subjects and patients were cultured with phytohemagglutinin stimulation, and IL-4 and IFN-γ concentrations were determined in the supernatant by enzymed linked immunosorbent assays. The percentage of Th1 and Th2, and the ratios of Th1:Th2 were correlated with cytokine (IFN-γ and IL-4) secretion level. These results demonstrated that Th2 cells were predominant in the second and third trimesters of normal pregnancy, but Th1 cells predominated in preeclamptic patients.
Mechanism of complement-dependent haemolysis via the lectin pathway: role of the complement regulatory proteins
Authors: Suankratay C , Mold C , Zhang Y , Lint T F , Gewurz H
Published: March 2003

Mannan-binding lectin (MBL) is an acute phase protein which activates the classical complement pathway at the level of C4 and C2 via two novel serine proteases homologous to C1r and C1s. We recently reported that haemolysis via this lectin pathway requires alternative pathway amplification. The present experiments sought to establish the basis for this requirement, and hence focused on the activity and regulation of the C3 convertases. Complement activation was normalized between the lectin and classical pathways such that identical amounts of bound C4 and of haemolytically active C4,2 sites were present on the indicator cells. Under these conditions, there was markedly less haemolysis, associated with markedly less C3 and C5 deposited, via the lectin pathway than via the classical pathway, particularly when alternative pathway recruitment was blocked by depletion of factor D. Lectin pathway activation was associated with enhanced binding in the presence of MBL of complement control proteins C4bp and factor H to C4b and C3b, respectively, with decreased stability of the C3-converting enzyme C4b,2a attributable to C4bp. Immunodepletion of C4bp and/or factor H increased lectin pathway haemolysis and allowed lysis to occur in absence of the alternative pathway. Thus, the lectin pathway of humans is particularly susceptible to the regulatory effects of C4bp and factor H, due at least in part to MBL enhancement of C4bp binding to C4b and factor H binding to C3b.
Effects of cocaine administration to influenza virus-immunized mice on cytokine profiles of individual splenic CD4+ and CD8+ T cells
Authors: Di Francesco P , Falchetti R , Gaziano R , Lanzilli G , Casalinuovo I A , Ravagnan G , Garaci E
Published: March 2003

We have analysed the effects of cocaine, administered to mice during the in vivo differentiation of effector T cells stimulated by antigen (influenza virus) recognition, on the frequency of IL-2-, IL-4- and interferon-gamma (IFN-γ)-expressing CD4+ and CD8+ T cells. Each animal was injected intraperitoneally with 10 mg/kg of cocaine 6, 24, 48 and 72 h after immunization with A/PR8 influenza virus (PR8). This enabled the determination of the pharmacological effects of cocaine on T cells during the initial step of the immune response, which is characterized by the production of large amounts of immunoregulatory cytokines. The distribution of IL-2-, IL-4- and IFN-γ-producing CD4+ and CD8+ T cells was assayed on unseparated PR8-immune spleen cells, obtained from mice treated with cocaine or vehicle, and restimulated in vitro with UV-inactivated PR8 virus. The frequency of T cells singly or co-expressing the above three cytokines was determined at single-cell level by simultaneous flow cytometric analysis of intracellular cytokines and surface antigen expression. In parallel, the levels of IL-2, IL-4 and IFN-γ in the culture supernatants were quantified by ELISA. The results showed that cocaine, administered during the in vivo virus-induced differentiation of T cells, caused an increase of both the frequencies of CD8+ T cells singly and co-expressing IL-2 and IFN-γ and the levels of these cytokines in virus-restimulated spleen cell culture supernatants, compared with those of untreated controls. In contrast, no effect was found on IL-4-positive CD8+ T cells and on IL-2-, IFN-γ- and IL-4-positive CD4+ T cells. Our findings suggest that the immunomodulatory effects of cocaine may be due to the up-regulation of the production of IL-2 and IFN-γ by CD8+ T cells with a type 0 cytokine profile.
Highly active anti-retroviral therapy (HAART) is associated with a lower level of CD4+ T cell apoptosis in HIV-infected patients
Authors: Roger P-M , Breittmayer J-P , Arlotto C , Pugliese P , Pradier C , Bernard-Pomier G , Dellamonica P , Bernard A
Published: March 2003

HAART may increase CD4+ T cell counts despite a persistently detectable HIV load. The impact of HAART on apoptosis, which may play a role in the disease process in HIV-infected patients, has not been extensively studied. We performed a study to compare the level of spontaneous T cell apoptosis and anti-retroviral treatments in a cohort of HIV-1-infected patients. Data were obtained from a computerized medical record. Quantification of apoptotic cells was by cytofluorometric technique. From November 1995 to December 1997 we studied T cell apoptosis in 112 HIV-infected patients. Forty patients were classified A, 36 B and 36 C. Thirty patients were naive and 82 received an anti-retroviral treatment, 49 including a protease inhibitor (PI). The median plasma viraemia determined in 63 patients was 3.6 (range 1.3–5.6) log10. The median apoptotic cell count was 22% (range 2–73%) and 12% (range 2–60%) for CD4+ and CD8+ T cells, respectively. We did not observe any correlation between the HIV viraemia and the level of apoptosis of T cell subsets. Patients with HAART showed a lower percentage of apoptotic CD4+ T cells only: 16% (range 2–61%) versus 25% (range 5–73%) for patients receiving two nucleoside analogues (P = 0.02). This effect was significant in stage A patients and remained observable during the whole course of HIV disease. In conclusion, HAART, without any relation to plasma viraemia, is able to reduce apoptosis of CD4+ T cells.
β2-adrenergic receptor gene polymorphisms in myasthenia gravis (MG)
Authors: Xu B-Y , Huang D , Pirskanen R , Lefvert A K
Published: March 2003

The β2-adrenergic receptor (β2-AR) belongs to the group of G-protein coupled receptors and is present mainly on skeletal and cardiac muscle cells and lymphocytes. The gene encoding β2-AR (ADRB2) displays a moderate degree of heterogeneity in the human population. The distribution of polymorphisms at amino acid positions 16, 27 and 164 is changed in asthma, hypertension and obesity. We have earlier reported a decreased density of the β2-AR on peripheral blood mononuclear cells and the presence of β2-AR antibodies in patients with MG. Since certain polymorphisms affect the function of the β2-AR, it was of interest to analyse these in MG. Using allele-specific polymerase chain reaction amplification, we revealed an over-representation of homozygosity for Arg16 and a lower prevalence of homozygosity for Gly16 in MG patients compared with healthy individuals. The increased frequency of homozygosity for Arg16 was due to a contribution from patients with generalized MG but not from patients with only ocular disease. Homozygosity for Glu27 was negatively associated with both the presence of β2-AR antibodies and severity of disease. Moreover, acetylcholine receptor (AChR) antibodies were more often present in patients being homozygous for Gln27. Our results imply that homozygosity for Arg16 confers susceptibility to generalized MG, and that certain polymorphisms at amino acid position 27 are associated with subgroups of patients.
Characterization of autoantibodies to endothelial cells in systemic sclerosis (SSc): association with pulmonary fibrosis
Authors: Ihn H , Sato S , Fujimoto M , Igarashi A , Yazawa N , Kubo M , Kikuchi K , Takehara K , Tamaki K
Published: March 2003

To determine the prevalence and the characterization of antibodies to endothelial cells in patients with SSc, serum samples from 80 patients with SSc, 20 patients with systemic lupus erythematosus (SLE), and 20 healthy control subjects were examined by ELISA using cultured human umbilical vein endothelial cells (HUVEC), indirect immunofluorescence analysis (IIF), and immunoblotting using cytoplasmic extract of HUVEC. IgG and/or IgM isotype anti-endothelial cell antibodies (AECA) were demonstrated by ELISA in 43 of 80 patients with SSc (54%), in 15 of 20 patients with SLE (75%), and in none of 20 healthy control subjects. Immunofluorescence analysis on HUVEC substrate showed homogeneous cytoplasmic staining. Immunoblotting demonstrated that these patients had antibodies directed to one or several antigens of approximately 60, 90, 110 and 140 kD, and the most common responses were to the 90-kD antigen. By the immunofluorescence method using HUVEC, affinity-purified anti-90-kD antibodies showed identical cytoplasmic staining to that produced by sera positive for AECA. Furthermore, AECA were closely correlated with pulmonary fibrosis in patients with SSc. These findings suggest that patients with SSc have abnormal antibodies to endothelial cell antigens, and support the hypothesis that endothelial dysfunction is involved in the development of this disease.
The thalidomide analogue CC-3052 inhibits HIV-1 and tumour necrosis factor-alpha (TNF-α) expression in acutely and chronically infected cellsin vitro
Authors: La Maestra L , Zaninoni A , Marriott J B , Lazzarin A , Dalgleish A G , Barcellini W
Published: March 2003

We investigated the in vitro effect of the water-soluble, highly stable thalidomide analogue CC-3052 on HIV-1 expression and TNF-α production in latently infected promonocytic U1 cells, acutely infected T cells and monocyte-derived human macrophages (MDM), and in mitogen-stimulated ex vivo cultures from patients with primary acute HIV-1 infection. HIV-1 expression was assessed by Northern blot analysis of RNAs, and ELISA for p24 antigen release and reverse transcriptase (RT) activity. TNF-α expression was evaluated by RT-polymerase chain reaction (PCR)-ELISA for mRNA and ELISA for protein secretion. We demonstrated that CC-3052 is able to inhibit HIV-1 expression, as evaluated by mRNA, p24 release and RT activity, in phorbol myristate acetate (PMA)- and cytokine-stimulated U1 cells. Furthermore, CC-3052 inhibited HIV-1 expression, as evaluated by p24 and RT activity, in acutely infected MDM and T cells. As far as TNF-α is concerned, CC-3052 significantly reduced TNF-α mRNA and protein secretion in PMA-stimulated U937 and U1 cells, and in PMA-stimulated uninfected and acutely infected MDM. Consistently, the addition of CC-3052 reduced TNF-α production in phytohaemagglutinin (PHA) and lipopolysaccharide (LPS)-stimulated whole blood cultures from patients during the primary acute phase of HIV-1 infection. Since TNF-α is among the most potent enhancers of HIV-1 expression, the effect of CC-3052 on TNF-α may account for its inhibitory activity on HIV-1 expression. Given the well documented immunopathological role of TNF-α and its correlation with viral load, advanced disease and poor prognosis, CC-3052 could be an interesting drug for the design of therapeutic strategies in association with anti-retroviral agents.
Neutrophil FcγRIIIb allelic polymorphism in anti-neutrophil cytoplasmic antibody (ANCA)-positive systemic vasculitis
Authors: Tse W Y , Abadeh S , Jefferis R , Savage C O S , Adu D
Published: March 2003

Neutrophils constitutively express FcγRIIa and FcγRIIIb receptors. Both receptors exhibit allelic variants which have different quantitative functional capacities: the biallelic FcγRIIa-R131 and -H131 alleles, and the neutrophil antigen (NA) NA1/NA2 alleles. ANCA activation of neutrophils requires ligation of FcγRIIa receptor, but recent data have shown that ANCA can also bind FcγRIIIb receptor. The aim of this study was to determine whether the FcγRIIIb polymorphism was a risk factor for the development of ANCA-associated systemic vasculitis, or the associated nephritis. FcγRIIIb receptor genotyping was determined by allele-specific polymerase chain reaction. Genomic DNA was extracted from 101 Caucasian patients with ANCA+ vasculitis (of whom 84 had renal disease) and 100 ethnically matched controls. Of the patients with ANCA+ systemic vasculitis, 71 had ANCA with specificity for proteinase 3 and 30 with specificity for myeloperoxidase (MPO). Overall no significant difference in genotype distribution or allele frequencies was found between patients and controls, or between patients with renal disease and controls. However, there was a trend for an increase in homozygosity for the NA1 allele in patients with a vasculitis and this was significant in patients who had anti-MPO antibodies. The FcγRIIIb receptor polymorphism is not a major factor predisposing to the development of ANCA+ systemic vasculitis or the associated nephritis. The over-representation of the FcγRIIIb homozygous NA1 allele in patients with anti-MPO antibodies may have implications for disease susceptibility.