Prolactin receptor heterogeneity in bovine fetal and maternal tissues

Recombinant human interferon-gamma (rhIFN-gamma) decreases the frequency of serious infections in patients with chronic granulomatous disease (CGD) through an unknown mechanism. To test the hypothesis that it exerts a beneficial effect by enhancing clearance of microbes from the bloodstream and tissues, normal human subjects were treated in vivo with rhIFN-gamma. Phagocyte opsonic receptor expression, serum opsonin levels, and phagocytosis of bacteria were then measured. A 4.7-fold increase in neutrophil expression of the high-affinity Fc gamma-receptor (Fc gammaRI) was observed that peaked 48 hours after the initiation of rhIFN-gamma treatment (P < .05). Monocyte expression of Fc gammaRI, Fc gammaRII, Fc gammaRIII, CD11a, CD11b, CD18, and HLA-DR also significantly increased with peak expression at 48 hours. Phagocytosis by neutrophils of killed Staphylococcus aureus opsonized with heat-inactivated pooled human serum significantly improved after rhIFN-gamma treatment (P < .05) and correlated with Fc gammaRI expression by neutrophils (r = .8, P < .001). This increase in ingestion could be inhibited by anti-Fc gammaRI monoclonal antibodies. Levels of the serum opsonin lipopolysaccharide-binding protein also significantly increased after in vivo rhIFN-gamma (P < .05). These results suggest that the protective effect of rhIFN-gamma in patients with CGD may involve improved microbial clearance. Moreover, improved phagocyte trafficking may occur secondary to increased expression of monocyte beta2-integrins. Because these IFN-gamma-related improvements in host defense were seen in normal hosts, rhIFN-gamma may have broader applications in the treatment of various disorders of immunity in addition to its demonstrated efficacy in CGD.     

Immunophenotyping of human bone marrow-derived macrophages

In vitro cultured cells derived from human bone marrow stimulated with macrophage-colony stimulating factor (M-CSF) or granulocyte-macrophage-colony stimulating factor (GM-CSF) were investigated for their immunophenotypic surface characteristics by flow cytometry. Approximately 80-90% of the cells showed morphological and histochemical features of macrophages and bore CD11a, CD11b, CD11c, CD14, CD29, CD32, CD33, CD44, CD45, CD54, CD64, CD71, HLA-DR antigen. The expression of CD4, CD25 and CD45RO were detected only in low density. Bone marrow-derived macrophages were negative for CD8, CD45RA, CD16 and CD23. Functionally, macrophages were able to phagocytose opsonized Escherichia coli, but no oxidative bursts were induced either by fmlp or by opsonized bacteria. Cell surface phenotyping revealed a CD pattern very similar to monocyte-derived phagocytes, and was partially distinct from resident stromal macrophages.     

Gross periostitis ossificans in mandibular osteomyelitis. Review of the English literature and radiographic variation

BACKGROUND: Although the airway epithelium participates in inflammation and repair, the circadian expression of epithelial cell markers involved in these processes has not been investigated. OBJECTIVE: We sought to determine whether expression of CD51 (vitronectin and fibronectin receptor), CD54 (intercellular adhesion molecule-1), HLA-DR (activation marker), CD29 (beta1 integrin), CD49b (collagen receptor), and CD11b (complement receptor) exhibit a circadian rhythm in asthma. METHODS: Eleven patients with nocturnal asthma (NA), 9 subjects with nonnocturnal asthma (NNA), and 10 control subjects underwent bronchoscopy at 4 PM and 4 AM in a random order 1 week apart, with brushing of the proximal and distal airways. The percentage of cells staining for a particular marker was determined. RESULTS: At 4 PM, HLA-DR in the proximal airways and CD54 in the distal airways was significantly greater in control subjects as compared with asthmatic subjects (HLA-DR, control subjects: 10.0% [range, 5.0% to 21.0%]; NNA: 8.0% [range, 4.0% to 14.5%] NA: 3.5% [range, 2.0% to 6.0%], P = .01; CD54, control subjects: 17.0% [range, 8.0% to 25.0%], NNA: 8.0% [range, 5.3% to 11.5%], NA: 7.0% [range, 4.0% to 15.0%], P = .O;). At 4 AM, CD51 in the distal airways was significantly greater in patients with NA as compared with patients with NNA and control subjects (control subjects, 23.0% [range, 13.8% to 30.5%]; NNA, 32.0% [range, 13.0% to 35.0%]; NA, 40.0% [range, 23.0% to 50.0%], P = .05). Expression of CD51 in the distal airways correlated with the degree of airway obstruction (r = -0.57, P = .001). Control subjects exhibited significant circadian variation in the expression of HLA-DR in the proximal airways and CD54 in the distal airways. CONCLUSION: The increased CD51 at night in patients with NA may be related to increased airway inflammation and repair processes in response to injury. The circadian changes in CD54 and HLA-DR in control subjects require further study to determine their significance. (J Allergy Clin     

Effects of r-metHuG-CSF on polymorphonuclear leucocyte kinetics and function in patients on continuous ambulatory peritoneal dialysis

End-stage renal failure (ESRF) patients undergoing continuous ambulatory peritoneal dialysis (CAPD) are immunocompromised and exhibit abnormal circulating polymorphonuclear leucocyte (PMN) function, including reduced phagocytosis and intracellular killing. Six uraemic patients on CAPD were each given 300 microg granulocyte colony stimulating factor (G-CSF) every day for 5 d and PMN function tests were performed daily. By day 5 of the study CD11b expression was significantly decreased in response to N-formylmethionylleucylphenylalanine (fMLP) and opsonized Staphylococcus epidermidis stimulation, and expression of L-selectin (CD62L) was significantly decreased in response to opsonized Staphylococcus epidermidis stimulation. Further, superoxide anion production and Fc gammaRI (CD64) expression were found to be significantly increased and Fc gammaRII (CD16) expression was lowered. Circulating white cell and PMN counts were significantly elevated in response to treatment. Administration of G-CSF did not appear to have corrected the abnormalities in phagocytosis and intracellular killing. This study suggests that G-CSF does no harm to ESRF patients and influences uraemic PMN function in a manner that is comparable to its effects on PMN in non-uraemic subjects.     

Phagocytosis, killing, and oxidant production by bovine monocyte-derived macrophages upon exposure to Brucella abortus strain 2308

Phagocytosis and intracellular survival of Brucella abortus, and oxidant production by monocyte-derived macrophages from ten B. abortus-naive cows were studied. Phagocytosis of bacteria opsonized with naive-autologous sera or reactor serum was significantly less than phagocytosis of bacteria opsonized with fetal bovine serum. After phagocytosis, intracellular survival of bacteria opsonized with naive-autologous or reactor sera was significantly less than survival of bacteria opsonized with fetal bovine serum. Production of oxidant by macrophages stimulated with B. abortus opsonized with naive-autologous, reactor, or fetal bovine sera was not significantly different. Although macrophages from one animal showed significantly less phagocytic activity, intracellular killing and oxidant production by macrophages from the ten individual cows toward B. abortus opsonized with naive-autologous, reactor, and fetal calf sera were homogeneous. The abilities of the macrophages to phagocytize and to kill B. abortus were not associated with each other or with oxidant production. Innate resistance or sensitivity to B. abortus was not identified in the cows based on macrophage function.     

Detection of in vivo proteasome activity in a starfish oocyte using membrane-impermeant substrate

The effect of calcitriol/1 alpha,25-dihydroxyvitamin D3, alone and in combination with cytokines, on the expression of various antigens (Ag) on human peripheral blood monocytes and U937 cells was studied by flow cytometry. Both constitutive and interferon-gamma (IFN-gamma), interleukin-4 (IL-4), IL-6 and tumour necrosis factor-alpha (TNF-alpha)-induced human leucocyte antigen (HLA)-DR, HLA-DP and HLA-DQ Ag expression on monocytes was significantly down-regulated by calcitriol, IL-10 and transforming growth factor-beta (TGF-beta). The effects of calcitriol were concentration dependent and reached maximal inhibitory levels after 3-5 days. Modulation of HLA-DR by calcitriol and IFN-gamma at the protein level correlated with the amount of mRNA specific for the HLA-DR alpha-chain, as judged by Northern blot analysis. The basal as well as IL-4, IL-6, IFN-gamma, TNF-alpha and TGF-beta-driven levels of HLA-ABC Ag were significantly diminished by calcitriol. On U937 cells calcitriol markedly induced CD11a and CD11b expression and weakly up-regulated CD11c whereas on monocytes, constitutive CD11a, CD11b and CD11c expression was significantly down-regulated by calcitriol. The expression of CD14 Ag was strongly induced on U937 cells but only modestly on monocytes. Both the basal level of CD71 and IL-4, IFN-gamma or TNF-alpha-driven expression was diminished on calcitriol-treated U937 cells. In addition, calcitriol suppressed the expression of CD71 Ag on monocytes. The ability of monocytes to phagocytize opsonized Escherichia coli was diminished by calcitriol. Our results demonstrate that calcitriol, alone or in combination with cytokines, modulates expression of MHC, CD11b, CD11c, CD14 and CD71 Ag on both monocytes and U937 cells, and impairs the phagocytic property of monocytes.     

Analysis of air pollution particulate-mediated oxidant stress in alveolar macrophages

Adverse health effects of urban air pollution particulates may be attributable to particle-mediated oxidant stress and inflammation. Intracellular oxidant production in normal hamster alveolar macrophages (AMs) was measured upon exposure to concentrated ambient particulates (CAPs), residual oil fly ash (ROFA), and their water-soluble and particulate fractions. ROFA and CAPs caused increases in dichlorofluorescin (DCFH) oxidation, a fluorescent measure of intracellular reactive oxygen species (ROS) production, comparable to the positive control, phorbol myristate acetate (PMA). The water-soluble component of both CAPs and ROFA (CAPs, S and ROFA, S) significantly increased AM oxidant production over negative control. CAPs samples and components showed substantial day-to-day variability in their oxidant effects. Metal chelation by desferrioxamine (DF, 1 mM) caused significant inhibition of particulate-induced AM oxidant production. ROFA exposure resulted in increased macrophage inflammatory protein-2 (MIP-2) message in AMs and in increased tumor necrosis factor alpha (TNF-alpha) production by the monocyte-macrophage cell line, RAW 264.7. TNF-alpha production was inhibitable by the antioxidant N-acetylcysteine (NAC). The data suggest that metal components adsorbed to urban air pollution particulates can significantly contribute to particulate ability to cause oxidant stress and cytokine production in AMs.     

Induction of death (CD95/FAS), activation and adhesion (CD54) molecules on blast cells of acute myelogenous leukemias by TNF-alpha and IFN-gamma

Leukemic growth is determined by the balance of cell proliferation, differentiation and cell death. In vitro, the blasts of acute myelogenous leukemia (AML) proliferate under the influence of certain positive and negative regulators (cytokines). We conducted this study to determine whether cytokines could induce markers of cell death (FAS/Apo-1/CD95), of cell activation (HLA-DR) and cell adhesion (ICAM-1, CD54) in AML cell lines and primary AML samples. As inducers, tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma were chosen. At baseline, CD95 and CD54 were weakly and HLA-DR was strongly expressed. CD95 was induced by TNF in 6/12 myeloid leukemia cell lines, and by IFN in 9/12 cell lines. Taken together, CD95 was upregulated by at least one cytokine in 11/12 cell lines. HLA-DR was inducible in 10/12 cell lines, with IFN being more potent than TNF. CD54 showed the strongest induction: TNF resulted in a more than 20-fold induction in positive cell lines, and IFN resulted in a more than 20-fold induction. In primary AML samples, CD95 was induced in 14/14 samples examined, with TNF being more potent than IFN. HLA-DR expression was increased by IFN in 12/15 samples and by TNF in 11/13 samples. The inducibility of HLA-DR by IFN was inversely correlated with baseline expression. As in the cell lines, CD54 was induced in most cases of AML. In addition to the induction of surface markers by cytokines, the culture of leukemia cells with fetal calf serum increased the expression of these markers, especially CD95 and CD54. Our results demonstrate that CD95 is not downregulated when TNF binds to its receptors, but is induced in cell lines and patient samples. Despite the induction of expression of CD95 (all cases of AML and most cell lines), 7/8 myelogenous leukemia lines and 6/7 patient samples remained resistant to CD95 triggering by antibody or by CD95 ligand, which suggests a lesion in normal cell signaling. As a positive control, a T-cell line (Jurkat) with 60% to > 90% apoptotic cells after a 22 h incubation was used. The number of CD95-binding sites was not correlated with the induction of apoptosis. The resistance of most cases of AML to CD95 triggering despite inducible expression may also be related to leukemia-specific antagonists of CD95 signal transduction, and requires further investigation. Altogether, our results indicate that surface markers related to apoptosis, activation and adhesion can be induced on AML blasts, and could be relevant to treatment strategies that exploit ligand binding to these surface epitopes.     

Infinity and the limits of the unconscious

Macrophages play an important role in the intestinal mucosal immune system. However, they are a poorly defined cell population. We therefore determined their phenotype in normal colonic mucosa. Macrophages were isolated from colonic biopsies and surgical specimens by collagenase digestion. Colonic macrophages were positively sorted by anti-CD33 magnetic beads. Flow cytometric triple fluorescence analysis was applied to study CD14, CD16, CD33, CD44, CD11b, CD11c, CD64, HLA-DR, CD80, CD86 and CD3/CD19 expression. CD33 was evaluated as a positive marker for intestinal macrophages. CD33+ cells isolated from normal colonic mucosa showed co-expression of the established intracellular macrophage marker CD68 in FACS analysis. CD33+ cells were capable of phagocytosis. Isolation of this cell population by magnetic anti-CD33 beads and culture resulted in a 4.2-40-fold increase in IL-1beta and 4.5-44-fold increase in tumour necrosis factor-alpha (TNF-alpha) secretion compared with unsorted lamina propria mononuclear cells (LPMC). Of the CD33+ cells, 90.9 +/- 6.9% (mean +/- s.d.) were CD44+. However, macrophages from colonic mucosa showed only a low expression of CD14 (10.5 +/- 3.8%), CD16 (10.1 +/- 3.9%), HLA-DR (27.3 +/- 9.2%), CD11b (17.4 +/- 6.8%), CD11c (17.8 +/- 10.4%). Furthermore, expression of CD80 (9.2 +/- 4.2%) and CD86 (15.1 +/- 7.3%) was low, suggesting a low ability of normal intestinal macrophages to activate T cells and T cell-mediated immune responses. We conclude that CD33 is useful for the isolation and flow cytometric characterization of colonic macrophages. These cells exhibit a single phenotype in normal mucosa (CD33++, CD44++, CD14-, CD16-, CD11b-, CD11c-, HLA-DRlow, CD80-, CD86-) lacking lipopolysaccharide (LPS) receptor and costimulatory molecules.     

Activation antigen expression on human T cells. I. Analysis by two-colour flow cytometry of umbilical cord blood, adult blood and lymphoid tissue

Activation antigens (actags) were detected on T cells at low levels of intensity by carefully defining negative cells with a panel of control antibodies. The mean percentage of blood T cells from healthy volunteers that expressed actags were 22% (CD25), 54% (CD26), 38% (CD38), 12% (CD54), 6% (CD69) and 21% (HLA-DR). The variability of actag expression detected by this sensitive method was determined on healthy volunteers by repeated estimation over a year. The percentage of T cells expressing CD25 and CD26 varied no more than repeated estimation of the CD4 T cell subset, whereas other actags showed greater variability. The antigen density of these actags on T cells was determined in relation to CD4 antigen density, and for most actags ranged from 10% to 75% of the level of CD4 antigen density except for CD7 and HLA-DR, which could exceed that of CD4. Different degrees of actag expression characterized T cells from different blood and lymphoid tissues. CD26, CD38 and CD45RA were universally expressed in cord blood at higher antigen density than adult blood. This immature pattern was consistent with recent thymic emigration. CD25, CD45RO, CD54 and HLA-DR progressively increased from cord blood through adult blood to lymphoid tissues, consistent with antigen-driven activation, whereas CD26 and CD45RA decreased. CD69, a very early activation antigen, abruptly increased in lymphoid tissue, exceeding CD25 by two-to-three-fold and suggesting a pre-activation state that may not involve commitment to antigen-driven proliferation. CD7 and CD38 expression was higher in cord blood and lymphoid tissue than in adult blood, indicating both an antigen-independent and -dependent up-regulation.     

Organization of the human RH50A gene (RHAG) and evolution of base composition of the RH gene family

The alpha4 chain (CD49d), which constitutes one of the chains of alpha4beta1 (very late activating antigen-4 [VLA-4]) and alpha4beta7 integrins, mediates migration of T cells to extravascular spaces. The interaction between VLA-4 and vascular cell adhesion molecule-1 (VCAM-1) has been shown to be the critical pathway for the selective accumulation of eosinophils and basophils at sites of allergic inflammation. T lymphocytes are also specifically recruited into allergic sites, including the allergic asthmatic airway. Increased numbers of activated CD4+ cells expressing the DR antigen subset of the human leukocyte antigens (HLA-DR) appear in the allergic lung 48 h after allergen inhalation. The mechanisms by which these cells localize into the lung are still unknown. We report that stimulation of allergen-specific T cells with allergen in vitro resulted in enhanced expression of alpha4 chain (CD49d) as measured by receptor density on allergen-specific T-cell lines and T-cell clones. Kinetic studies showed that CD49d density was enhanced over a 24- to 48-h period in a time-dependent fashion, and was coordinately upregulated with HLA-DR expression. We also demonstrated that increased expression of CD49d on T-cell lines 24 h and 48 h after stimulation correlated with increased adhesion to the CS-1 fragment of fibronectin. In contrast, lymphocyte function-associated antigen-1b (LFA-1b) (CD11b), LFA-3 (CD58), and intercellular adhesion molecule-1 (ICAM-1) (CD54) expression did not change with allergen stimulation. We also showed that CD49d receptor density on T cells obtained by bronchoalveolar lavage (BAL) of allergic patients before and 48 h after allergen challenge was significantly higher than that on T cells taken from BAL of normal subjects and from controls with other inflammatory lung diseases. Taken together, these findings indicate that allergen stimulation activates allergen-specific T cells and coordinately induces increased CD49d receptor expression and binding to counterligands. We postulate that allergen-driven upregulation of CD49d, which together with the beta1 chain constitutes VLA-4 integrin, may be responsible for the selective accumulation of T cells in the allergic asthmatic lung.     

Selective alterations in binding kinetic parameters and allosteric regulation of N-methyl-D-aspartate receptors after prolonged seizures in the developing rat brain

To investigate the effect of chronic smoke exposure on pulmonary macrophages (PM), the expression of seven different surface and intracellular molecules of PM was studied in induced sputum (IS) samples from healthy volunteers--nine smokers and seven non-smokers. Sputum was induced by inhalation of nebulized saline (3.5% NaCl). Cell viability and total cell counts (TCC) were performed immediately. Cell differentials were determined on May-Grunwald Giemsa-stained cytospin preparations. The PM were immunologically characterized by use of the following monoclonal antibodies: RFD1, RFD7, CD11b, CD54, CD68, CD71 and HLA-DR. The stainings were performed with a three-step, indirect immuno-alkaline phosphate method. Viability and TCC did not differ between the groups. Smokers had a higher percentage of macrophages (P < 0.05) and a lower proportion of neutrophils (P < 0.05). The percentage of macrophages expressing RFD1, HLA-DR, CD71 (P < 0.01 for all) and CD54 (P < 0.05) was significantly lower in smokers, whereas the remaining markers were expressed equally in the two groups. The results indicate that smoking induces a decrease in the expression by PM of surface molecules known to be associated with the antigen-presenting function.     

Humoral immunity and regulation of intrapulmonary growth of Legionella pneumophila in the immunocompetent host

The potential role of humoral immunity in regulating intrapulmonary growth of Legionella pneumophila in the immunocompetent host was investigated using a murine model of Legionnaires' disease. Intratracheal inoculation of A/J mice with a virulent strain of L. pneumophila (10(6) bacteria per mouse) resulted in the recruitment of B lymphocytes into the lung and the development of anti-L. pneumophila Ab. Opsonization of L. pneumophila in vitro with anti-L. pneumophila-specific mAb resulted in a significant decrease in intrapulmonary growth of the bacteria at 24 to 72 h postinfection. Transmission electron microscopic analysis of lung tissue from L. pneumophila- infected mice demonstrated that while there was no significant difference between phagocytosis of the unopsonized and opsonized L. pneumophila by alveolar macrophages at 24 h postinfection, phagocytosis of opsonized bacteria by alveolar mononuclear phagocytic cells was significantly enhanced at 48 h postinfection. Depletion of A/J mice of complement before intratracheal inoculation of opsonized L. pneumophila (10(6) bacteria per mouse) did not significantly alter intrapulmonary growth of L. pneumophila. These results suggest that anti-L. pneumophila Ab, produced during replicative L. pneumophila lung infections, may regulate intrapulmonary growth of L. pneumophila in the immunocompetent host by decreasing the viability of extracellular L. pneumophila and by enhancing phagocytosis of the bacteria by alveolar mononuclear phagocytic cells by a complement-independent mechanism.     

Searching for Lyme disease in Colombia: a preliminary study on the vector

Since ageing is accompanied by various alterations of the immune system, the aim of the present study was to investigate the effect of age on the expression of surface markers in peripheral blood monocytes. We studied 28 healthy young subjects and 28 elderly subjects fulfilling the criteria of the SENIEUR protocol. Peripheral blood mononuclear cells were isolated and the expression of various surface markers was analysed by double colour flow cytometry. The mean fluorescence intensity of the intercellular adhesion molecule- (CD54), CD29, CD45RO and CD32 was increased significantly in CD14+ cells of elderly people when compared with the young subjects. No significant differences were found in the expression of CD11a, CD11b, CD15, CD26, CD27, CD33, CD45RA, CD45RB, CD49d, HLA-DR and CD65. In summary, we demonstrated significant increase in four monocyte surface markers in subjects of old age; this finding may be related to certain immune phenomena observed in ageing subjects such as auto-immunity.     

The interrelation of the expression of HLA class I and II proteins and of the costimulatory proteins CD11a/CD18 and CD16 with the lytic activity of resting and activated natural killer lymphocytes

It has been presumed that: 1) rheumatoid arthritis (RA) is a genuine model of activation of the peripheral natural killers (NK); 2) methods of NK separation can cause their activation, and therefore only peripheral mononuclear cells were used. The aim of this research was to study the role of cytolysis of HLA-I, HLA-DR, and costimulatory proteins CD11a and CD16 expressed on resting and activated peripheral NK cells. A total of 74 healthy volunteers (HV) and 74 RA patients were compared before corticosteroid or gold treatment. Two-colour immunofluorescence and cytofluorimetric analysis, cytotoxic test against the target K 562 cells (class I negative) before and after incubation of PMC with monoclonal antibodies (MoAB) to HLA-I (E3D8), HLA-DR (ICO-1), CD11 (ICO-11), PXx63 Ag8.653 as control, and also incubation with recombinant alpha 2-IFN were used. VEP-13 MoAB was used for CD16 determination. Compared with HV PMC, the RA PMC showed a statistically significant increase of activated NK lymphocytes (CD11a+DR+ and CD16+DR+ by 2.03 and 1.96 times, resp.), and increased level of expression of HLA-I, HLA-DR, and CD16 on CD11a+ lymphocytes, resulting in significantly diminished cytolytic activity (CA) by 1.9 times. A short-term incubation with alpha 2-IFN (100 U/ml, 1 h) increased the level of expression of HLA- and costimulatory proteins and the CA of the HV PMC, while the RA PMC did not react. HV peripheral NK are concluded to be mainly resting cells, and the RA peripheral NK are mainly the activated ones. Only using the level of expression of HLA-DR on CD11a lymphocytes, as a ranking criterion, allowed to reveal two types of correlation between the CA and immunocytological parameters of PMC. The first type: in HV, the CA correlated positively (P < 0.025) with the percentage of CD11a+CD16+, and with levels of expression of HLA-I (P < 0.025) and CD16 (P < 0.025) expressed on CD11a lymphocytes (5-9.5 MFI DR--the mean fluorescence intensity of HLA-DR). The second type: the CA positively correlated (P < 0.05) with the percentage of CD11a+CD16-, and with levels of expression of HLA-DR (P < 0.05) and CD11a (P < 0.05) expressed on CD11a lymphocytes (12.4-16 MFI DR). In RA patients with a "normal" density expression of HLA-DR on CD11a lymphocytes (up to 14 MFI) there exists a mixed type: the CA correlated positively (P < 0.05) with the percentage of CD11a+CD16+, and with levels of expression of HLA-DR (P < 0.05) and CD11a (P < 0.05) expressed on CD11a lymphocytes. In RA patients with a very high density expression of HLA-DR on CD11a lymphocytes (VH-DR group, 18-26 MFI) the CA correlated negatively with the percentage of CD11a+CD1 6+ (P < 0.05), and with levels of expression of HLA-DR (P < 0.025) and CD11a (P < 0.025) expressed on CD11a lymphocytes. The anti-HLA-I,-DR, -CD11a MoABs strongly inhibit the Ca in HV- and RA-PMC. However, in the VH-DR group the anti-DR MoAB stimulates the CA from 27.06 +/- 10.25 up to 41.69 +/- 14.23%. Thus, activated peripheral CD11a+CD16+ lymphocytes suppress the CA of other NK.     

Effect of storage and ultraviolet B irradiation on CD14-bearing antigen-presenting cells (monocytes) in platelet concentrates

BACKGROUND: Ultraviolet B (UVB) irradiation of platelet concentrate (PCs) reduces platelet alloimmunization, but the mechanism of the effect is unclear. Evidence suggests that UVB may downregulate the expression of surface adhesion molecules on passenger antigen-presenting cells in PCs. STUDY DESIGN AND METHODS: The effect of blood bank storage, platelet preparation from whole blood, and UVB irradiation on the quantitative expression of intercellular adhesion molecule-1 (ICAM-1, or CD54), HLA-DR, CD45, and CD11c on CD14-positive antigen-presenting cells (monocytes) was studied by using two-color flow cytometry. RESULTS: Blood bank storage for 4 days resulted in upregulation of ICAM-1 and HLA-DR and downregulation of CD14 but left the expression of CD11c and CD45 unchanged. Preparation of PCs from fresh whole blood was associated with a rapid increase in CD11c without upregulation of ICAM-1 and HLA-DR. UVB irradiation before storage inhibited the upregulation of ICAM-1 and HLA-DR, resulted in accelerated downregulation of CD14, and was associated with increased loss of monocytes. Agitation of the PC bag during irradiation was of critical importance, since omission of agitation resulted in largely uninhibited upregulation of ICAM-1 but was still associated with significantly higher cell loss than that seen in unirradiated controls. CONCLUSION: UVB exposure nonspecifically affects monocytes in PCs, resulting in downregulation of surface molecules that are important for antigen presentation, as well as in significant cell loss.     

Macrophage-parasite relationship in theileriosis. Reversible phenotypic and functional dedifferentiation of macrophages infected with Theileria annulata

Theileria annulata is a tick-transmitted protozoan parasite of cattle, which transforms cells of macrophage (Mphi) or B cell lineage. Bone marrow cells, bone marrow cell-derived, and monocyte-derived Mphi were infected with T. annulata sporozoites, and the resulting cell lines were assessed for surface marker expression and function. Transformed lines expressed histocompatibility complex (MHC) class-I and II, CD44, CD45, and the myeloid marker DH598-surface markers CD14, CD11b, M-M7, TH57A, and to a lesser extent CD11a/CD18, CD11c, and ACT(B), were down-regulated. Likewise, transformed cells failed to express Mphi functions (Fc-receptor-mediated phagocytosis, phorbol myristate acetate-induced oxidative burst, lipopolysaccharide-induced tumor necrosis factor alpha, and nitric oxide generation and procoagulant activity up-regulation). Mphi origin was assured by homogeneity of the starting population, cloning of cells by limiting dilution, and repeated microscopic and flow cytometric monitoring of the cell lines. Elimination of the parasite by treatment with BW720c resulted in the re-acquisition of monocyte lineage properties, as evidenced by up-regulation of CD14, and by re-acquisition of the capacity to ingest opsonized sheep red blood cells and bacteria. Thus, Mphi transformed by T. annulata appear to undergo a process of parasite-induced dedifferentiation but reassume the differentiated phenotype upon elimination of the parasite.     

Effects of monocyte purification and culture on integrin expression

Selective alterations in the surface expression of members of the LeuCAM (leukocyte cell adhesion molecule) family of integrins occur during in vitro culture of human monocytes. Such changes may relate in part to cellular maturation, but also to activation following purification and culture of monocytes. In this paper, we examined the effects of monocyte isolation, adherence during culture and endotoxin exposure on the expression of these molecules and the ligand for LFA-1, ICAM-1 (CD54). Expressions of CD11b, CD18 and CD54, but not CD11a or CD11c, were higher on monocytes freshly isolated by density gradient separation and plastic adherence as compared with cells labelled directly in whole blood. However, the surface expression of the LeuCAMs and CD54 on cultured monocytes was not affected by short-term adherence to plastic for 2 h, as determined by comparisons of their expression on adherence-isolated and elutriated monocytes. In contrast, prolonged adhesion of monocytes for up to 21 days in culture altered expression of CD11a without affecting that of the other LeuCAMs or CD54. Expression of CD11a decreased more rapidly on adherence-maintained cells as compared with suspension-cultured cells. Our results show that cellular manipulations required for in vitro studies of monocyte/macrophages may alter expression of the LeuCAMs.     

The NF-kappa B inhibitor, tepoxalin, suppresses surface expression of the cell adhesion molecules CD62E, CD11b/CD18 and CD106

Tepoxalin, a dual enzyme inhibitor of cyclooxygenase and 5-lipoxygenase has been shown to inhibit T-cell activation. Its immunosuppressive property is distinct from cyclosporin because only tepoxalin, but not cyclosporin, suppresses NF-kappa B activation. Here we report that tepoxalin selectively inhibits intercellular adhesion molecule-1 (ICAM-1, CD54)/MAC-1 (CD11b/CD18) dependent adhesion of polymorphonuclear cells to IL-1 activated human umbilical vein endothelial cells. The mechanism of inhibition is related to the surface expression of several cell adhesion molecules. Flow cytometry analyses on cultured cells that were treated with tepoxalin or antisense oligonucleotides to the P65/p50 subunit of NF-kappa B, and then stimulated with PMA, revealed a reduced expression of CD11b/CD18 on monocytic HL60 cells, and endothelial adhesion molecule-1 (CD62E) and vascular adhesion molecule-1 (CD106) on human umbilical vein endothelial cells. Expression of other adhesion molecules such as lymphocyte function associated-antigen-1 (CD11a/CD18) and CD54 were unaffected. Tepoxalin also inhibited the secretion of a NF-kappa B regulated chemokine, IL-8, a known inducer of CD11b/CD18 expression. Thus the suppression of CD11b/CD18 expression by tepoxalin may involve IL-8. Our results suggest that by inhibiting NF-kappa B activation, surface expression of several adhesion molecules can be modulated and that tepoxalin may be useful in treating selected adhesion mediated events such as leukocyte migration or atherosclerotic plaque formation.