bluuebunny/crossref_metadata_2025 huggingface dataset:
The Involvement of Kupffer Cells in the Clearance of High Molecular Weight Rat IgA Aggregates in Rats
Authors: BOGERS W. M. J. M. , GORTER A. , JANSSEN D. J. , RITS M. , BAZIN H. , VAN ES L. A. , DAHA M. R.
Published: June 2006

In the present study the clearance kinetics and tissue distribution of aggregated 125I‐labclled monoclonal rat IgA ([125I] AIgA) of different sizes were studied in rats. Soluble [125I]AIgA disappeared from the circulation in a biphasic manner with an initial rapid distribution half‐life (TI) and a second slower half‐life (T2). T2 was directly related to the size of the aggregates. High molecular weight [125I]AIgA, containing 10–12 IgA molecules per aggregate ([lgA]10–12). was cleared much faster than low molecular weight aggregates. The main site of clearance was the liver. The larger the size of the AIgA, the more degradation products were found in the circulation. After injection of [[lgA]10–12, non‐parenchymal cells (NPC) contained three times more radioactivity than parenchymal cells(PC)(NPC:P ratio 3.06 ± 0.96). Ratios of 0.82 ± 0.0.3 and 0.62 ± 0.12 were observed when [IgA]5–6 and [IgA]2 were injected respectively, suggesting a greater role for Kupffer cells in the clearance of large‐sized IgA aggregates. Kupffer cells were shown to be the main cells for localization of large‐sized AIgA established by immunohistochemical staining on liver cryostat sections.
Accessory Signals in T‐T Cell Interactions Between Antigen‐ and Alloantigen‐Specific, Human Memory T Cells Generated in Vitro
Authors: ØDUM N. , RYDER L. P. , GEORGSEN J. , JAKOBSEN B. K. , GEISLER C. , MøLLER J. , MORLING N. , DlCKMEISS E. , SVEJGAARD A.
Published: June 2006

The potential of activaied HLA class II‐positive T cells as anligen‐/alloantigen‐presenting cells remains controversial. In our model system we use in vitro‐primed. HLA class II‐specitic T cells of the memory T‐cell phenotype, CD4+. CD29+ (4B4+), and CD45RO+ (UCHL‐1), We have previously shown that alloactivated. HLA class II‐posilive T cells (Ta) are unable to stimulate proliferalive responses in naive and primed allospecific T cells when ‘back‐stimulation’ is avoided. The explanation of this feature of Ta is unknown, but it is due neither to suppression nor to insufficient HLA class II expression. Accordingly, we investigated the possibility that Ta have a deficient expression of accessory signals critical for the induction of proliferative T‐cell responses. We found that (I) non‐mitogenic concentrations of phorbol myristate acetale (PMA) in combination with either rIL‐4, a CD28‐reaclive MoAb (Kolt‐2). or a calcium ionophore (A23187) enabled Ta to elicit alloantigen‐specific memory T‐cell responses and to present purified protein derivative (PPD) to PPD‐speciftc T‐cell lines. The addition of irradiated, Epstein–Barr virus‐transformed B‐cell lines (EBV‐LCL) (but not their supernatants) had a similar but less pronounced effcet; (2) MoAb directed against HLA class II, CD25 (IL‐2R), CD2, CD4, CD11a (LFA‐1), or CD45RO molecules inhibited these responses; (3) PMA was required within the first hour of culture in order to induce optimal alloantigen‐specifie T‐cell aetivation. while rIL‐4 was fully effective when added after 20–44 h of culture; (4) incubation for 20 h of Ta with rIL‐4 plus PMA markedly up‐regulated CD54 expression on the Ta, and IL‐4 seemed to potentiate the effect of PMA on the CD54 expression. In conclussion, the present data indicate that the inability of Ta to elicit (allo)antigen‐speciffe, proliferative T‐cell response is due to a lack of critical accessory signals. Up‐regulation of CD54 was not sufficient for Ta to stimulate proliferative responses. Neither cytokines (IL‐1, IL‐6, and others) nor triggering of CD2 epitopes (T11.2 and T11.3) by soluble MoAb or solid phase support by MoAb against a number of accessory molecules provided the necessary signals. Thus, our data indicate that other, as yet unknown, signalling pathways play a key role in antigen‐ and alloantigen‐specitic T–T interactions. These palhways still need to be identified.
Intradermal Recombinant Interleukin 2 Enhances Peripheral Blood T‐Cell Responses to Mitogen and Antigens in Patients with Lepromatous Leprosy
Authors: CONVERSE P. , OTTENHOFF T. H. M. , TEKLEMARIAM SABA WORK , HANCOCK G. E. , DIETZ M. , BECX‐BLEUMINK M. , WONDIMU ASSEFA , KIESSLING R. , COHN Z. A. , KAPLAN G.
Published: June 2006

Thirty‐one patients with lepromatous leprosy received recombinant interleukin 2 (IL‐2) intradermally in doses ranging from 10 to 30 μg. Before injection and at time intervals of 2–21 days thereafter, samples of peripheral blood mononuclear cells (PBMC) were obtained. Single or multiple injections (1–3) of IL‐2 did not modify the total number of circulating lymphocytes or the number of T cells and the CD4/CD8 T‐cell ratio However. IL‐2 had a pronounced influence on the [3H]thymidine incorporation in response to various stimuli 4–8 days alter intradermal IL‐2. Stimulation indices of three‐ to sevenfold above pre‐IL‐2 levels were observed with the polyclonal activator phytohaemagglutinin (PHA) and enhanced thymidine incorporation occurred in the presence of antigens to which the patients were already sensitized, such as purified protein derivative and BCG. IL‐2 had no effect on the unresponsive stale of lepromatous leprosy patient T cells to the antigens of Mycobacterium leprae.
The Pathogenesis of Leishmania aethiopica Infection in BALB/c Mice
Authors: AKUFFO H. O. , WALFORD C. , NILSEN R.
Published: June 2006

A mouse model for L. aethiopica infection is described. BALB/c mice were unable to clear an infection with 1 × 107 promastigotes injected into the hind footpad. However, there was no ulceration of the lesion and no development of overt clinical symptoms after 203 days of infection. Spread of viable organisms was evident in the draining lymph node but not in the spleen or liver. The control of the infection was associated with the development of classical delayed hypersensitivity responses to phenolized promastigotes and appeared as a localized granulomatous infiltration. The infiltration had features of classical tuberculoid granulomas, but superimposed on it was a strong eosinophilic infiltration. The relevance of such cells though unclear is discussed.
Specificity of Two Subsets of Cytotoxic Human γδ T‐Cell Clones
Authors: QVIGSTAD E. , BOSNES V. , LUNDIN K. E. A. , THORSBY E.
Published: June 2006

Peripheral blood mononuclear cells were enriched for γδ T cells by immunomagnetic separation, stimulated with cells from an allogeneic donor, and cloned. T‐lymphocyte clones (TLC) of the two major γδ T‐cell subsets, BB3− (i.e. Vδ2+)and δTCSI+ (i.e. Vδ1/(D)/Jδ1). were obtained. In addition, one γδ TLC was BB3−δTCSI+. All of the BB3+ TLC showed strong cytotoxicity against various allogeneic tumour cell lines, such as Daudi and K562. The cytotoxicity against the tumour cell lines was modulated by MoAb against the γδ TcR. The BB3+ TLC were not cytotoxic against B‐lymphoblastoid cell lines (B‐LCL). In contrast, the δTCSI+ TLC showed much lower cytotoxic activity against the tumour cell lines, but many were strongly cytotoxic against allogeneic B‐LCL. Some of the δTCSI + TLC demonstrated HLA‐specific cytotoxicity, while other δTCSI+ TLC had a more broad cytolytic activity against B‐LCL. Thus, the two major subtypes of γδ T cells from this donor, as defined by MoAb BB3 and δTCSI, were distinct with respect to recognition specificity.
Concomitant Effect of 2′‐Deoxycoformycin on Natural Killer Cell Activity and Tumour Cell Sensitivity to Lysis in Hairy Cell Leukaemia—Discordant Effects of Alpha Interferon
Authors: LAHAT N. , AGHAI E. , MERCHAV S. , KINARTY A. , SOBEL E. , FROOM P.
Published: June 2006

The effects of 2′‐deoxycoformycin (dCF) and alpha interferon (IFN‐a) on natural killer (NK) cell‐enriched fractions and hairy cell (HC) targets from three patients with HC leukaemia (HCL) were investigated. There was no significant increase in NK activity when either the HC targets or NK‐enriched cells were preincubated with dCF. However, preincubation of both HC and NK cells with dCF resulted in increased NK activity. Culture of enriched NK cells with IFN‐α enhanced their activity. However, preincubation of HC targets with this drug in the presence or absence of dCF resulted in a protective effect. Maximal NK activity towards HCL was obtained when the target tumour cells were separately precultured with dCF and the NK‐enriched effectors precultured with dCF + IFN‐α. The effect of dCF and IFN‐α was also measured using the standard K562 cells as targets for NK activity. dCF enhanced NK activity following preculture of both effector and target K562 cells, but IFN‐a did not reduce K562 cell susceptibility to NK lysis as it did for HC cells. Our findings suggest that (a) dCF and IFN‐α, which are used to treat HC, could function via activation of NK cells, (b) effects on both effector and tumour target cells should be taken into account, and (c) caution should be exercised in extrapolating the effects of NK‐cell activity against K562 cells to those on HC targets.
Interleukin 1, but not Interleukin 1 Inhibitor, is Released from Human Monocytes by Immune Complexes
Authors: REMVIG L. , THOMSEN B. S. , BAFK L. , SVENSON M. , BENDTZEN K.
Published: June 2006

Immune complexes (IC) are believed to play a role in the pathogenesis of some autoimmune diseases in which interleukin 1 (IL‐1) and probably other cytokines also take part. This investigation shows that tetanus toxoid human anti‐tetanus toxoid IC induce human monocytes to release IL‐1. The activity was identified as being mainly IL‐1β by molecular size chromatography, isoelectric focusing, and anti‐IL‐1β affinity chromatography. Endotoxins were eliminated by repetitive washing of the IC suspension and by preincubation of IC with polymyxin B. The IL‐1‐inducing effect of IC was destroyed by healing at 80° C. and it was not blocked by the cytoskeleton inhibitor cytochalasin B. IL‐1 inhibitors were not detected in the supernatants.
Monocyte Function in IDDM Patients and Healthy Individuals
Authors: MØLVIG J. , POCIOT F. , BÆK L. , WORSAAE H. , WOGENSEN L. DALL , CHRISTENSEN P. , STAUB‐NIELSEN L. , MANDRUP‐POULSEN T. , MANOGUE K. , NERUP J.
Published: June 2006

Interleukin 1β(IL‐1β) and tumour necrosis factor alpha (TNF‐α) may be pathogenetically important in insulin‐dependent diabetes mellitus (IDDM), which is associated with genes of the HLA region. Since a regulatory role of HLA region genes on monokine production may exist, we looked for an association between the monokine and prostaglandin E2 (PGE2) responses of monocytes (Mo) from 20 healthy males (18–50 years) with HLA‐DR types relevant for IDDM susceptibility and resistance (DR 1,2, DR 1,3, DR 1,4, DR 3,4). Monokine assays were established and evaluated and the secretions of IL‐1β, TNF‐α, and PGE2 measured in Mo cultures (2 h, 6 h, 20 h) prepared by endotoxin‐free techniques and stimulated by low‐dose E. coli lipopolysaccharides (LPS). There were no significant associations between Mo responses and HLA‐DR phenotype. Likewise, Mo from DR2 (n=5) and DR4 (n= 5) homozygous healthy males demonstrated no significant differences in monokine and PGE2 responses of Mo.

In the HLA class III region a diallelic TNF‐β gene Ncol polymorphism consisting of alleles of 5.5 kb and 10.5 kb was recently described and associated with susceptibility to autoimmune diseases including IDDM. We report that IL‐1β and TNF‐α responses of Mo from TNF‐β 10.5 kb homozygous healthy individuals were significantly higher than for TNF‐β 5.5/10.5 kb heterozygotes.

IL‐1β and TNF‐α responses of Mo from males (18–35 years) with newly diagnosed (n= 10) and long‐standing IDDM (n= 10) and from age‐ and HLA‐DR‐matched healthy males (n= 10) were studied. LPS, gamma interferon (IFN), and TNF‐α‐stimulated Mo cultures were investigated. No significant differences were found between Mo responses of IDDM patients and controls. IFN (1000 U/ml) in the presence of LPS significantly potentiated LPS‐stimulated Mo TNF‐α secretion and reduced the levels of IL‐β immunoreactivity in Mo lysates. IFN and TNF‐α did not have any effects on LPS‐stimulated Mo secretion of IL‐1 β immunoreactivity.

We conclude that Mo IL‐1β and TNF‐α production is normal in patients with recent‐onset and long‐standing IDDM. The interindividual differences in monokine responses may be accounted for by the diallelic human TNF‐β gene polymorphism rather than by HLA class II genes. This observation may be important for understanding the association of certain H LA haplotypes with autoimmune phenomena and disease.
Dynamics of Blood Components and Peritoneal Fluid during Treatment of Murine E. coli Sepsis with β‐1, 3‐D‐Polyglucose Derivatives
Authors: RASMUSSEN L.‐T. , SELJELID R.
Published: June 2006

Beta‐1, 3‐D‐polyglueose derivatives protect mice against otherwise lethal bacterial infections This protective effect has previously been considered to be mediated through mononuclear phagocytes. We have now investigated the cellular composition in blood and peritoneal fluid after administration of the β‐1,3‐D‐polyglucose before and after challenge with Escherichia coli. In animals treated with β‐1,3‐D‐polyglucose derivatives, the total white cell number was significantly increased in both blood and peritoneal fluid before and after challenge with E. coli. The increased total cell number was mainly the result of raised levels of granulocytes. The effects of β‐1,3‐D‐polyglucose‐derivatized microbeads (GDM) and soluble aminated β‐1,3‐D‐polyglucose (AG) were similar.

Bacterial counts in peripheral blood in GDM‐ and AG‐treated animals increased until 6 h after challenge and approached zero after 24 h. In untreated animals the bacterial counts increased gradually until the animals died after about 12 h. Bacterial counts in peritoneal fluid of GDM‐and AG‐treated animals declined to zero after 24 h. In untreated animals there was a slight increase in bacterial counts until the animals died after about 12 h.

By using radioactive labelling, we localized the bacteria as well as the β‐1,3‐D‐polyglueosc derivatives during the period following injection. Particle‐bound β‐1,3‐D‐polyglucose was recovered mainly in the milky spots of the omentum. A conspicuous number of bacteria were also recovered in the milky spots. The soluble aminated β‐1,3‐D‐polyglucose was recovered mainly in the liver. However, on a weight basis, the greatest concentration of radioactivity was in the milky spots.
Cardiac Amyloidosis: Report of a Patient Heterozygous for the Transthyretin Isoleucine 122 Variant
Authors: SARAIVA M. J. M. , SHERMAN W. , MARBOE C. , FIGUEIRA A. , COSTA P. , DE FREITAS A. F. , GAWINOWICZ M. A.
Published: June 2006

Amyloid fibril material was extracted from autopsy material of a patient who died from progressive cardiac failure at age 64. He had an enlarged heart on routine X‐ray at age 47 and the first symptoms of cardiac failure at age 62. Fractionation of the fibril material resolved peptide fragments immunoreactive with anti‐human transthyretin (TTR). One of the peptides was farther purified by high‐performance liquid chromatography (HPLC) and subjected to tryptic peptide mapping along with TTR isolated from the patient's serum. In both instances, sequencing procedures revealed, in addition to the normal peptide 12 (residues 105 126). an abnormal peptide with an isoleucine for valine substitution at position 122. This substitution has been described previously in a patient with systemic senile amyloidosis (SSA) homozygous for this variant. The results question whether SSA is a clinical entity related to TTR He 122 with phenotypic expression in the homozygous condition.
Suppression of Target Cell Proliferation by Natural Killer Cells
Authors: JÚLÍUSSON P. B. , ÖGMUNDSDÓTTIR H. M.
Published: June 2006

The cytolytic effects of natural killer (NK) cells have been extensively studied in recent years. In the present study we have investigated the cytostatic effects of SK cells Human peripheral blood lymphocytes from healthy volunteers were used as a source of effector cells, and the cell lines K562, U937, U1285, and Molt‐4 were used as target cells Effector cells were enriched for NK cells Using Percoll gradients and depleted of NK cells on Percoll gradients or by using Leu‐19 antibodies and magnetic heads. By monitoring cell numbers during co‐culture of effector cells and K562, it was found that after an initial phase of cell killing for 3 h target cell numbers remained stable during the following 24‐48h. In a microcytotoxicity assay measuring inhibition of uptake of [3H]thymidine, the four target cell types were shown to have different NK sensitivity; inhibition of ≥ 80% was obtained for K562 and U937 at an effector to target cell (E/T) ratio of 30:1, 50% for U1285, and 30% for Molt‐4. This inhibition was shown to be partly a direct effect on DNA synthesis for all cell lines, as Incorporation of [3H]thymidine was decreased in co‐cultured target cells compared with an equal number of target cells alone. Inhibition of DNA synthesis was thus not directly related to cell death and was also observed for the Molt‐4 cell line‐that was not killed. A cell division assay, with target cells m agarose and effector cells m a liquid tippet layer, showed a decline in the rate of target cell divisions Effects on the cell cycle were studied on latent‐phase cells. It was shown that effector cells delayed the onset of DNA synthesis This anti‐proliferative effect was observed for several days, but cell growth then gradually resumed the effector cells were identified as CD56‐positive large granular lymphocytes (I. GL). Double‐layer cultures and experiments using effector cell supernatants demonstrated that the growth‐inhibitory effect could be mediated by soluble factors, and the production of such factors was stimulated by exposure to a small proportion of target cells (50:1), Studies with specific antibodies indicated that growth inhibition was not mediated by alpha interferon (IFN‐α) but it was partly mediated by tumour necrosis factor alpha (TNF‐α). It is concluded that NK cells have a growth‐inhibitory effect that is distinct from the cytolytic effect and this activity is probably mediated by several soluble factors including TNF‐α.
Analysis of Age‐Related Degeneracy of T‐Cell Repertoire: Localized Functional Failure in CD8+ T Cells
Authors: KOMURO T. , SANO K. , ASANO Y. , TADA T.
Published: June 2006

The repertoire and frequency of alloreactive T cells in aged mice were examined by limiting dilution analysis of the mixed‐lymphocyte reaction (MLR). Mice were aged in specific pathogen‐Tree conditions up to 31 months, and MLR of their spleen cells under limiting dilution conditions was examined at various stages of ageing. The Frequency of alloreactive T cells was found to decrease in mice more than 14 months of age, and the value reached about 1/3 of that of young mice of 2 months of age. The slope analysis of cell dose response curves of MLR indicated the existence of two interacting T‐cell populations, i.e. CD4+ and CD8+, cooperation between which is required for the full manifestation of MLR in limiting dilution conditions. The decrease in the alloreactivity of aged spleen cells was ascribed to the selective functional loss of the CD8+ population, because the CD4+ hut not the CD8+ T‐cell traction from aged mice could restore the limiting factor in MLR. The decrease in the cooperative activity of CD8+ T cells in allo‐MLR starts much earlier than the decrease m the number of CD8+ T cells in the spleen.
Tγδ Cells and their Subsets in Blood and Synovial Tissue from Rheumatoid Arthritis Patients
Authors: SMITH M. D. , BRÖKER B. , MORETTA L , CICCONE E. , GROSSI C. E. , EDWARDS J. C. W. , YÜKSEL F. , COLACO B. , WORMAN C. , MACKENZIE L. , KINNE R. , WESELOH G. , GLÜCKERT K. , LYDYARD P. M.
Published: June 2006

We have examined the frequencies of Tγδ cells in blood, synovial fluids, and synovial membranes of patients with rheumatoid arthritis (RA) and in blood from age‐matched controls Immunocyto‐chemical and immunohistochemical techniques were used with monoclonal antibodies BB3 and A13 to define a major and minor blood subset of Tγδ cells respectively. Together, these antibodies identify the majority (if not all) of the peripheral blood Tγδ cells.

Significantly lower levels of Tγδ cells were found in the blood of RA patients compared with controls, whilst higher but not significant numbers were found in the synovial fluids or paired samples Scattered Tγδ cells were found only in some synovial membranes with A distribution similar to the Tγδ cells Analysis of the two different Tγδ ‐cell subsets indicated a ratio of BB3 to A13 of about 5:1 in control and RA blood. However, this ratio was less than 1:1 in the RA synovial fluids and membranes. The migratory nature of the A13+ cells could account for their predominance in these sites the possible pathological significance of these cells in the rheumatoid synovial fluid and synovial membranes is discussed.
Control of the Expression of a Class II Major Histocompatibility Gene (HLA‐DR) in Various Human Cell Types: Down‐Regulation by IL‐1 but not by IL‐6, Prostaglandin E2, or Glucocorticoids
Authors: WATANABE Y. , LEE S. , ALLISON A. C.
Published: June 2006

We have examined the effects of recombinant human IL‐1α and IL‐1β on expression of the gene for the class II major histocompatibility antigen, HLA‐DR, and the class I MHC antigen, HLA‐B7. induced by natural or recombinant human IFN–γ in several human cell types. Recombinant hIL‐1α and hIL‐1β antagonized the class II MHC‐inducing effect of IFN‐γ in human monocytes and normal skin fibroblasts, and in chondrosarcoma and astrocytoma cell lines. In the presence of IL‐1α or IL‐1β, the induction by IFN‐γ of HLA‐DR mRNA. and the expression of the corresponding antigen on the cell surface, were reduced when analysed by dot‐blot hybridization and flow cytometry respectively. Nuclear transcription assays showed that IFN‐γ augmented expression of the HLA‐DR gene and that IL‐1α inhibited this effect. IL‐I‐mediated inhibition was not observed with IFN‐γ‐induced expression of MHC class I mRNA (HLA‐B7). Both IL‐α and IL‐β induced production of IL‐6 mRNA and of PGE; in these cell types. However, recombinant IL‐6 or PGE2 did not inhibit IFN‐γ‐induced HLA‐DR mRNA expression. Dexamethasone did not inhibit HLA‐DR expression in the cells studied but eliminated the inhibitory effect of IL‐1 on such expression. The observations suggest that MHC gene expression is influenced by the cytokine network in a complex manner which is different for class I and II genes. Glucocorticoids and PGE2 do not consistently inhibit class II expression.
Oxidative Burst in Human Epstein‐Barr Virus‐Transformed B‐Cell Lines Triggered by Immobilized Specific Antigen
Authors: MALY F.‐E. , URWYLER A. , TENGLER R. , STEINITZ M. , GOLDING B. , DE WECK A. L.
Published: June 2006

Normal human B lymphocytes and Epstein‐Barr virus‐transformed B‐cell lines can produce reactive oxygen species such as superoxide if treated with phorbol myristate acetate (PMA) or with the surface immunoglobulin cross‐linking agents protein A and anti‐immunoglobulin. Here, we investigated under which conditions specific antigen, the natural ligand of surface immunoglobulin, can stimulate an oxidative burst in monoclonal Epstein Barr virus‐transformed B‐cell lines producing antibodies of known specificities. After a short lag time of 1‐2 min, exposure to the specific antigen stimulated a prolonged oxidative burst (tmax, 30‐90 min). as measured by Lucigenin‐enhanced, superoxide dismutase‐inhibitable chemiluminescence, in the corresponding line only. The effect was induced in each line if the specific antigen was immobilized to a solid support. Except in one line in which antigen also stimulated an oxidative burst if presented at relatively high density on a soluble carrier, soluble antigen did not induce B‐cell oxidase activation. This suggests that normal, non‐transformed B lymphocytes also require interaction with relatively dense deposits of specific antigen for activation of their oxidase.
Phagocytosis of Agarose Beads by Receptors for C3b (CR1) and iC3b (CR3) on Alveolar Macrophages from Patients with Sarcoidosis
Authors: PETTERSEN H. B. , JOHNSON E. , OSEN S. S.
Published: June 2006

Alveolar macrophages (AM) from sarcoidosis patients exhibit no detectable defect in their potential to synthesize the functional alternative and terminal pathway of complement. They also synthesize more C9 than AM from healthy controls. Various authors [4, 6] have suggested that sarcoid AM have decreased phagocytic ability. In the present work we studied whether there was any difference in C3 receptor‐mediated phagocytosis of serum‐treated and native agarose beads by AM recovered from patients with active sarcoidosis compared with controls, AM from seven patients with active sarcoidosis and seven healthy controls were cultured under serum‐free conditions for 2, 12, 24. and 48 h. We found a significantly increased CR1 and CR3 receptor‐mediated phagocytosis of native agarose heads by AM from the seven patients. CR1 and CR3 were also detected on AM directly recovered from bronchoalveolar lavage fluid using fluorescein‐conjugated monoclonal anti‐receptor antibodies. The percentage of AM expressing CR appeared to be increased in sarcoidosis. The reason for the enhanced phagocytosis of agarose beads by the sarcoid AM is probably the result of both increased synthesis and receptors of complement. Altered complement production and complement receptors may be important for the pathogenesis of this granulomatous disorder.
Immunological Marker Analysis of Mitogen‐Induced Proliferating Lymphocytes Using Brd U Incorporation or Screening of Metaphases
Authors: ADRIAANSEN H. J. , OSMAN C. , VAN DONGEN J. J. M. , BRFSSER J. H. F. M. WIJDENES‐DE , TILBORG C. M. J. M. KAPPETIJN‐VAN , HOOIJKAAS H.
Published: June 2006

The proliferative effects of the mitogens phylohaemagglutinin (PHA). concanavalin A (Con A), pokeweed mitogen (PWM). and staphylococcal protein A (SpA) were investigated using two different methods which enable immunological marker analysis of proliferating cells: either surface marker labelling followed by Brd U incorporation or screening of metaphases after surface marker labelling. Therefore peripheral blood mononuclear cells from six healthy volunteers were stimulated with these four mitogens. Both PHA and Con A gave rise to more CD8+ than CD4+ proliferating cells. PHA, but not Con A, induced B‐cell proliferation as well. PWM mainly caused T‐cell proliferation. SpA also appeared to be a potent T‐cell mitogen in addition to its capacity to induce B‐cell proliferation. However, in contrast to the other mitogens SpA predominantly stimulated CD4+ cells.
The Antibody Repertoire of Early Human B Cells
Authors: LYDYARD P. M. , QUARTEY‐PAPAFIO R. P. , BRÖKER B. M. , MACKENZIE L. , HAY F. C. , YOUINOU P. Y. , JEFFERIS R. , MAGEED R. A.
Published: June 2006

Our previous studies have shown that a high frequency of Epstein‐Barr virus (EBV)‐immortalized cord blood (CB) and fetal liver (FL) clones produce IgM antibodies which display extensive autoreactivity for IgG Fc (rheumatoid factor, RF). To investigate further the repertoire of these early B B cells we have examined the expression of CRI associated with RF paraproteins in relation to antibody specificity and polyreactivity. CRI were detected by ELISA and/or flow cytometry using a panel of well‐characterized monoclonal antibodies defining idiotopes associated with particular VKand VH gene family products and raised against Fc‐specific paraproteins.

Many of the CRI were expressed by these clones, suggesting that they may be markers of early B cells. The presence of the CRI was not always associated with Fc specificity. Three of eight CB/FL clones expressed the VKII subgroup of light chains, and two of these expressed the VK sub‐subgroup associated CRI, 17‐109. These two clones reacted with IgG Fc, and one also bound to single‐stranded DNA. The VHIII‐associated idiotope D12 was expressed on IgM from 4 out of 9 FL and 5 out of 12 CB clones. D12 and B6(also a VH‐III‐associated CRI) were coexpressed in 4 out of 5 CB clones but not in the four FL clones. Seven out of nine clones expressing these idiotopes were polyreactive. and five had Fc‐binding activity.

Three of the 12 CB clones expressed the VHI‐associated conformational idiotope G8. One of 20 CLL clones expressed both B6 and D12, and another expressed both 17‐109 and the VH‐I associated G6and G8 idiotopes. Taken together, these data provide evidence for the frequent usage, in early B cells, of VK subgroups and VH‐associated idiotopes of RF paraproteins. The expression of these CRI was not a prerequisite for binding to IgG Fc. but there was a frequent association of these idiotopes with it. Differences in expression of CRI between CLL and early B‐cell clones may suggest differences in the pattern or VH usage between these subsets of B cells.
T‐Cell Responses to Fractionated Antigens of Mycobacterium w, a Candidate Anti‐Leprosy Vaccine, in Leprosy Patients
Authors: YADAVA A. , SURESH N. R. , ZAHEER S. A. , TALWAR G. P. , MUKHERJEE R.
Published: June 2006

Mycobacterium w, an atypical cultivable mycobacterium. is undergoing phase III clinical trials as a vaccine against leprosy in India. It has brought about lepromin conversion and histopathological upgradation in a significant number of patients studied so far. It is important to identify antigens of W. w that trigger T‐cell responses in leprosy patients vaccinated with this organism. In the present study the peripheral T‐cell repertoire of 12 M. w‐vaccinated leprosy patients. 10 unimmunized leprosy patients. 8 tuberculoid and 5 healthy contacts was analysed with fractionated antigens of M. w. The lepromatous leprosy patients who are in general anergic to antigens of M. leprae did not respond to antigens of M. w. However, peripheral blood mononuclear cells obtained from leprosy patients who had been vaccinated with M. w responded to many antigens. These responses were frequently directed against low molecular weight entities of 14‐ 45 kDa. T cells from tuberculoid leprosy patients and healthy contacts also responded predominantly to a number of low molecular weight antigens of M. w. The study also identified an immunodominant 28‐31 kDa antigenic fraction carrying T‐ as well as B‐cell activating determinants.
Mobilization of Gelatinase‐Rich Granules as a Regulatory Mechanism of Early Functional Responses in Human Neutrophils
Authors: MOLLINEDO F. , PULIDO R. , LACAL P. M. , SANCHEZ‐MADRID F.
Published: June 2006

Components involved in superoxide anion production (cytochrome b) and in cell adhesion processes (CD11b, CD11c. CD18), two early functional responses of neutrophils during acute inflammation, are intracellularly located in resting human neutrophils. We have found a correlation between secretion of gelatinase and overexpression in the plasma membrane of CD11b. CD11c. CD18 and cytochrome b upon cell activation. Gelatinase and lactoferrin were parallely released after cell activation with different stimuli, but a better correlation between antigen up‐regulation and gelatinase release was obtained. Total translocation of the intracellular pool of these mobilizable molecules to plasma membrane was achieved under conditions that induced total degranulation of the gelatinase‐rich granule population, whereas 50% and 90% of the lactoferrin‐containing secondary granules and peroxidase‐containing primary granules, respectively, remained unfused. These results suggest a mechanism by which neutrophil function can be regulated through mobilization of gelatinase‐rich granules, which can be considered as a subpopulation of secondary granules.
Expression and Secretion of T‐Cell Receptor Vα and Vβ Domains using Escherichia coli as a Host
Authors: WARD E. S.
Published: June 2006

An expression system for the production of recombinant T‐cell receptor (TCR) variable domains would, inter alia, allow structural studies lo be carried out and provide protein for the generation of anti‐clonotypic antibodies, in this report the Vα and Vβ domain genes have been isolated from a T‐cell hybridoma which is associated with the pathogenesis of experimental allergic encephalomyelitis (EAE) In the H‐2 mouse. These have been expressed as secreted domains in Escherichia coli, using secretion vectors previously used for the production of immunoglobulin fragments. Both Vα and Vβ domains are secreted in milligram quantities into the culture supernatant, although the levels of the Vα domain are about 10‐20 Ibid higher than those of the Vβ domain. This expression system offers a rapid route for the production of recombinant TCRs in soluble form.
Mechanisms in Neutrophil Priming: Characterization of the Oxidative Response Induced by Formylmethionyl‐Leucyl‐Phenylalanine in Human Exudated Cells
Authors: FOLLIN P. , BRIHEIM G. , DAHLGREN C.
Published: June 2006

Exudate human polymorphonuclear neutrophils were isolated and investigated regarding oxidative responsiveness and priming ability. The exudate neutrophils were found to produce an increased amount of O2‐ and H2O2 when stimulated with formylmethionyl‐leucyl‐phenylalanine (fMLP). i.e. these cells were metabolically primed. Cytochalasin B (cyl B) pretreatment affected the production of O2‐ by exudate cells, although to a lesser extent than the production by peripheral blood cells, in which a substantial increase was induced. Addition of N‐ethylmaleimide (NEM) to activated exudate and peripheral blood cells revealed no difference in oxidase in activation rate. To induce further priming, the cells were incubated in vitro with a synthetic diacylglycerol (sn‐1,2‐didecanoylglycerol: diC10), or the Ca2+ ionophore ionomycin. Results of this procedure showed significant differences between exudate and peripheral blood neutrophils: the peripheral cells expressed a primed response, which was measured as increased fMLP‐induced O2‐ production following incubation with both these substance: where a the metabolic activity of exudated cells was not affected by diC10 but was significantly primed by ionomycin (P<0.01). The exact route for diacylglycerol priming is unknown. However, our results with human neutrophils primed during exudation indicate an exhausted diC10‐priming pathway, with a retained sensitivity for priming [Ca2+]i rises.
Presence of NAP‐1/IL‐8 in Synovial Fluids Indicates a Possible Pathogenic Role in Rheumatoid Arthritis
Authors: PEICHL P. , CESKA M. , EFFENBERGER F. , HABERHAUER G. , BROELL H. , LINDLEY I. J. D.
Published: June 2006

The synovial fluid in affected joints of rheumatoid arthritis (RA) patients contains many cells, in numbers strongly correlated with the severity of disease. As the disease worsens and the cell count increases, the polymorphonuclear leucocyte becomes the predominant cell type. Although the inflammatory cytokines interleukin 1 (IL‐I) and tumour necrosis factor (TNF) have no direct neutrophil‐attractant activity, they are both potent inducers of interleukin 8(IL‐8) in a variety of cell types, Chemotactic attraction of neutrophils is a major activity of IL‐8.

Examination of a number of synovial fluids showed that significant levels of IL‐8 art present in a high proportion of R A cases (10 out of 17). at concentrations directly related to the number of cells in the joint, and to circulating C‐reactive protein (CRP) levels. The cytokine is present only at background levels in other diseases accompanied by arthritic manifestations, including systemic lupus erythematosus (SLE) and induced arthritis. The progressive joint destruction seen in all cases where high IL‐8 levels were measured, coupled with the neutrophil‐rich cell count and the strong correlation between concentration of IL‐8 and both serum CRP and cellular influx into the joint, is strongly suggestive of a pathogenic role for IL‐S in RA.
The Human Acute‐Phase Serum Amyloid A Gene Family: Structure, Evolution and Expression in Hepatoma Cells
Authors: BETTS J. C. , EDBROOKE M. R. , THAKKER R. V. , WOO P.
Published: June 2006

Serum amyloid A (SAA) proteins are a group of phylogenetically conserved acute‐phase reactants. Evidence is presented here for the existence of four genetic loci for the human serum amyloid A (SAA) genes. The first locus was defined by three contiguous γ clones spanning =: 30 kb which contained a single SAA gene encoding apoSAA1β. Allelic variants were isolated at (he second locus: a novel clone encoding apoSAA2α was distinguished from SAA2β (previously known as SAAg9, Ref. 1) by a His/Arg polymorphism at residue 71. SAA 1 and SAA2 found in the high density lipoprotein fraction of acute‐phase plasma were =90% homologous at the nucleotide level. Homology in the 5’flanking regions was reflected functionally with similar transcriptional responses to inflammatory cytokines in transfected hepatoma cells. A further novel gene, SAA4. was isolated from a cosmid library and mapped 10 kb downstream of SAA2. The locus defining SAA3 has been described elsewhere [2]. Polymorphisms were detected at both SAA1 and SAA2 loci by Southern analysis and the entire SAA region mapped to discrete fragments by pulsed field analysis. The four genes account for all the hybridizing bands present on Southern analyses in a Caucasian population.
CD3‐Induced T‐Cell Proliferation and Interleukin‐2 Secretion is Modulated by the CD45 Antigen
Authors: ORAVECZ T. , MONOSTORI É. , KURUCZ É. , TAKÁCS L. , ANDÓ I.
Published: June 2006

In the present study we have investigated the effect of CD45, C D45RA and CD45RO monoclonal antibodies (MoAbs) on the CD3 receptor‐mediated proliferation of human T lymphocytes. It is shown that CD3‐induced proliferation of purified resting T cells and quiescent T lymphoblasts (QT L) is promoted via all of the investigated CD45‐associatcd epitopes. It is also shown that the CD45 molecules are required to be cross‐linked for costimulation. The MoAbs enhance the interleiikin‐2 (IL‐2) production of CD3‐stimulated QTL. The elevation of the IL‐2 production correlates with the increase in CD3‐inducod cell proliferation suggesting that the CD45‐driven regulation of T lymphocyte activation is linked to the IL‐2 pathway.
Expression of HLA Class I and II (DR, DP and DQ) Determinants in Fetal and Postnatal Salivary Glands
Authors: THRANE P. S. , HALSTENSEN T. S. , ROGNUM T. O. , BRANDTZAEG P.
Published: June 2006

Fetal (n=20)and postnatal (n = 40) parotid glands were examined by two‐colour immunohisto‐chemistry combining monoclonal and polyclonal antibody reagents lo study the expression of HLA class I and Il (DR. DP. and DQ). CD45 and CD3, The epithelium lacked class II during fetal life, whereas class I determinants appeared in some acini and most major ducts. Fetal vessels were positive for both class I and class II (mainly DR), suggesting constructive expression. Some class II‐positive (DR > DP DQ) histiocytic cells, scaltered CD45+ leucocytes, and very few CD3+ T cells were present in the fetal stroma. The epithelium remained DR‐negative the first few weeks after birth, but brisk expression was seen subsequently, DP and DQ remained virtually negative in the epithelium throughout the first year. A slight postnatal increase of class II expression (DR > DP > DQ). along with an apparent decrease in class I. was observed in the endothelium. The number of class Il‐positive histiocytic cells. CD45+ leucocytes and CD3+ T cells, as well as the proportion of presumably activated (DR+) T cells, increased a few weeks after birth. The local immune system hence seemed to be stimulated by extrinsic factors, but the overall number of T cells nevertheless remained small. Stimuli other than T cell‐derived lymphokines, therefore, probably explained the brisk postnatal epithelial DR induction.
The Pharmacokinetics, Distribution and Degradation of Human Recombinant Interleukin 1β in Normal Rats
Authors: REIMERS J. , WOGENSEN L. D. , WELINDER B. , HEJNÆS K. R. , POULSEN S. S. , NILSSON P. , NERUP J.
Published: June 2006

Based upon in vivo rat experiments it was recently suggested that interleukin I in the circulation may be implicated in the initial events of β‐cell destruction leading to insulin‐dependent diabetes mellitus (IDDM) in humans. The aim of the present study was to estimate half‐lives of distribution and elimination phases (T1/2β) of human recombinant interleukin 1β(rIL‐1β), and its tissue distribution and cellular localization by means of mono‐labelled, biologically active 125I‐rIL‐1β. After intravenous (iv.) injection, 125I‐rIL‐1β was eliminated from the circulation with a T1/2α of 2.9 min and a T1/2β of 41.1 min. The central and peripheral volume of distribution was 20.7 and 19.1 ml/rat, respectively, and the metabolic clearance rate was 16.9 ml/min/kg. The kidney and liver showed the highest accumulation of tracer, and autoradiography demonstrated that 125I‐rIL‐1β was localized 10 the proximal tubules in the kidney and to the hepatocytes in the liver. Furthermore, grains were localized to the islets of Langerhans in the pancreas. Tracer‐bound proteins corresponding to intact 125I‐rIL‐1β were found in the circulation after i.v., intraperitoneal (i,p.) and subcutaneous (s.c.) injections, as demonstrated by high performance size exclusion chromatography. trichloracetic acid precipitation and SDS PAGE until 5h after tracer injection. Pre‐treatment with “cold” rIL‐1β enhanced degradation of a subsequent injection of tracer. The route of administration was of importance for the biological effects of rIL‐1β as demonstrated by a reduced food intake, increased rectal temperature and blood glucose after s.c. injection of rIL‐1β compared with i.p. The present demonstration of intact rIL‐1β in the circulation and the islets of Langerhans supports the hypothesis that systemic IL‐1β may be involved in the initial 1β‐cell destruction leading to IDDM in humans.
Effects of Castration, Depo‐Testosterone and Cyproterone Acetate on Lymphocyte T Subsets in Mouse Thymus and Spleen
Authors: ABOUDKHIL S. , BUREAU J. P. , GARRELLY L. , VAGO P.
Published: June 2006

The effects of testosterone on the relative proportion of Thy 1.2, CD4 (L3T4) and CD8 (Lyt‐2) cells in thymus and spleen were studied after castration and administration of Depo‐testosterone (DT) separately or together with cyproterone acetate (CA) (an antiandrogen) in BDFI mice. Injection of 0.5 mg/100 g body weight of DT during 2 weeks decreased significantly the number and proportion of double positive (DP) (CD4+ CD8+) and increased the percentage of single positive (SP) CD4+ (CD4+ CD8‐), whereas there was a slight decrease in the Thy 1.2+ cells in the thymus. In parallel, we observed an increase in CD8+ (CD4‐ CD8+) cells in the spleen. The androgen deprivation after 3 weeks of castration induced a decrease in the percentage of CD4” cells in thymus and both CD4+ and CD8+ cells in spleen. Injection of CA (0.5 mg/100 g body weight) had the same qualitative effects as DT on the proportion of lymphocyte T subsets in castrated mice. However, the combined activities of DT and CA were greater than either alone. These data indicate the main role of testosterone in the distribution of CD4+ and CD8+ cells in male mice. The similar effects of CA and DT in the lymphoid organs may suggest a difference between androgen receptors of sexual and lymphoid organs.
Separation of Leucocytes: Improved Cell Purity by Fine Adjustments of Gradient Medium Density and Osmolality
Authors: BØYUM A. , LØVHAUG D. , TRESLAND L. , NORDLIE E. M.
Published: June 2006

This paper briefly reviews commonly used procedures for separation or mononuclear cells (MNC) and granulocytes from human blood with X‐ray contrast media as gradient material, and also presents new and modified procedures for leucocyte preparation. Standard techniques for human blood do not always yield satisfactory results with blood from other species. In general pure MNC are easily obtained (top fraction), but often the granulocyte fraction has a low purity, due to contamination with MNC that move to the bottom during centrifugation and contaminate the granulocyte suspension. Obviously the density distribution of MNC differs between species, However. the separation can be improved by tine adjustment of gradient medium osmolality. For this purpose we have used Nycodenz a non‐ionic X‐ray contrast medium, A favourable property of Nycodenz solutions is that the osmolality and density can easily be varied over a broad range. The cells react promptly to a change of medium osmolality. In hypertonic medium the cells expel water. shrink, their density increases and they sediment faster, in spite of a smaller radius. Further. the cells may pass what was initially a density barrier. A hypotonic environment has the opposite effect.

In the present work we were able to show that a slight change of medium osmolality clearly improved different techniques for separation of leucocyte subgroups. For instance, the Isopaque Ficoll (IF) technique consistently yielded MNC and granulocytes of high purity with human blood. However, with blood from rabbits, rats and mice the granulocyte suspensions were contaminated by 40‐60%, MNC‐ By utilizing Nycodenz. and lowering the osmolality by 10‐12 per cent (at constant density–1.077 g/ml) we obtained satisfactory separation of MNC as well as granulocytes with blood from these species.

A problem in the routine separation of granuloccytes (IF) is a high contamination of erythrocytes (2 5 per cell) in the granulocyte suspension. With a two‐layer technique with Nycodenz solutions of different densities it was possible lo separate granulocytes almost devoid of erythrocytes. after proper adjustment of osmolatily, By appropriate combination of density and osmolality, Nycodenz was a suitable gradient material in other separation procedures as well, e.g. the separation of monocytes and mast cells.

To facilitate the use of Nycodenz as a versatile gradient material, a computer program providing recipes for various Nycodenz solutions is included as an appendix.
Enhancement of IgE Synthesis in the Human Myeloma Cell Line U‐266 with an IgE Binding Factor from a Human T‐Cell line
Authors: NILSSON G. , JERNBERG H. , HELLMAN L , AHLSTEDT S. , NILSSON K.
Published: June 2006

An IgE‐binding factor(s) (IgE‐BF(s)) was partially purified from the supernatant of human HTLV‐II carrying T‐cell line MO, This IgE‐BF(s) was shown to increase the IgE synthesis in the human myeloma cell line U‐266, but did not affect its viability or growth. The of effect of the IgE‐BF(s) was dose‐dependent and selective for IgE protein synthesis as β2‐microglobulin synthesis in the U‐266 and the immunoglobulin production in the U‐1958 IgG‐secreting human myeloma cell line were unaffected. The IgE‐BF(s) increased the production of the c heavy chain but not the X light chain production. The IgE‐BF(s) was distinct from IL‐1β. IL‐3, IL‐4, IL‐5, lL‐6, TNF‐α, IFN‐α ‐β, ‐γ, M‐CSF, and fragments of CD23.
Rabbit Serum Amyloid Protein A: Expression and Primary Structure deduced from cDNA Sequences
Authors: RYGG M. , MARHAUG G. , HUSBY G. , DOWTON S. B.
Published: June 2006

Serum amyloid A protein (SAA), the precursor of amyloid protein A (A.A) in deposits of secondary amyloidosis. is an acute phase plasma apolipoprotein produced by hepatocytes. The primary structure of SAA demonstrates high interspecies homology. Several isoforms exist in individual species, probably with different amyloidogenic potential. The nucleotide sequences of two different rabbit scrum amyloid A cDNA clones have been analysed, one (corresponding to SAA1) 569 base pairs (bp) long and the other (corresponding to SAA2) 513 bp long. Their deduced amino acid sequences differ at five amino acid positions, four of which arc located in the NH;‐terminal region of the protein. The deduced amino acid sequence of SAA2 corresponds to rabbit protein AA previously described except for one amino acid in position 22. Eighteen hours after lurpentine stimulation, rabbit SAA m RNA is abundant in liver, while lower levels are present in spleen. None of the other extrahepatic organs studied showed any SAA mRNA expression, A third mRNA species (1.9 kb) hybridizing with a single‐stranded RNA probe transcribed from the rabbit SAA cDNA. was identified. SAA 1 and SAA2 mRNA were found in approximately equal amounts in turpentine‐stimulated rabbit liver, but seem to be coordinately decreased after repeated inflammatory stimulation.