bluuebunny/crossref_metadata_2025 huggingface dataset:
Modifications Induced by Activation to Tumor Cytotoxicity in the Protein Secretory Activity of Macrophages
Authors: Grand-Perret Thierry , Petit Jean-François , Lemaire Geneviève
Published: January 2018

After biosynthetic radiolabeling, polypeptides secreted by macrophages were analyzed by electrophoresis on polyacrylamide gels in the presence of sodium dodecyl sulfate and fluorography.

Comparative studies of various macrophage populations (inflammatory macrophages, primed and activated macrophages, resident macrophages) showed that secretory activity is elaborately controlled according to macrophage developmental and functional state.

When young macrophages activated to cytotoxicity against tumor cells by an appropriate eliciting agent and culture in the presence of lipopolysaccharide were compared to mature resident macrophages devoid of antitumor activity, the following changes were observed in the protein secretion phenotype: (1) New polypeptides are expressed: the intensity of four bands was increased more than five-fold on one-dimensional gels and 17 new spots were detected on two-dimensional gels. (2) The production of several major polypeptides is repressed: six bands were strongly decreased on one-dimensional gels, and 11 major spots (or groups of spots) were lost on two-dimensional gels. (3) The charge microheterogeneity pattern exhibited by several polypeptides on two-dimensional gels is shifted towards less acidic species.
Staphylococcus aureus-Derived Chemoattractant Activity for Human Monocytes
Authors: Rot Antal , Henderson Louis E , Leonard Edward J
Published: January 2018

Products of bacteria are potent chemoattractants for mammalian leukocytes. Several reports suggest that these attractants are small peptides. We compared the properties of culture fluids of Staphylococcus aureus with fMet-Leu-Phe, considered a prototype of bacterial attractant. Chemotactic activity for human monocytes of Staph. aureus culture filtrates was determined in multiwell chemotaxis chambers. At optimal concentrations, the filtrate attracted almost twice as many monocytes as fMet-Leu-Phe (53 ± 5% of input number compared with 30 ± 3%, in a series of ten experiments). Gel-filtration characteristics and susceptibility to proteolytic digestion suggested that chemotactic activity was due to peptides with a molecular size range of 500–2,000 daltons. Reverse-phase high-pressure liquid chromatography (HPLC) of unfractionated filtrate revealed nine peaks of chemotactic activity, most of which was in five of the peaks. One peak accounted for 40% of total activity. Individual peaks, like the unfractionated material, were capable of attracting about twice as many monocytes as the optimal concentration of fMet-Leu-Phe. Quantitative bioassay of the HPLC peaks showed that only 5% of the total Staph. aureus chemotactic activity eluted in the position of fMet-Leu-Phe. This is in contrast to the report that fMet-Leu-Phe accounted for 70% of neutrophil lysosomal enzyme-releasing activity of Escherichia coli culture fluid. In summary, chemotactic activity for monocytes of Staph. aureus culture fluid is due to peptides other than fMet-Leu-Phe; these peptides recruit a higher percentage of monocytes than fMet-Leu-Phe.
BCG, in Contrast to C. parvum, Does Not Induce Increased Sensitivity to the Toxic Effects of Indomethacin in Mice
Authors: Hart David A
Published: January 2018

Treatment of mice with killed Corynebacterium parvum (also designated Propionibacterium acnes, P. acnes) or Bacille Calmette Guerin (BCG) leads to modification of several of the same host systems. However, BCG, in contrast to C. parvum, did not induce increased sensitivity to the toxic effects of indomethacin in BALB/c or C57B1/6 mice. In addition, treatment of mice with BCG did not interfere with the induction of sensitivity by C. parvum. Therefore C. parvum must uniquely induce changes in host systems which alter the sensitivity to this anti-inflammatory drug. Additional experiments with splenectomized animals revealed that the presence of this organ, which undergoes hypertrophy following C. parvum treatment, was not necessary for the induction of indomethacin sensitivity. Presentation of C. parvum via the subcutaneous route versus the intraperitoneal route revealed that the two routes were equally efficient in inducing sensitivity in C57B1/6 mice but the former route was less effective (50% deaths) than the intraperitoneal route (95% deaths) in BALB/c mice. These results indicate that host related factors (genetic) may be important in the generation of enhanced sensitivity to the toxic effects of indomethacin.
Human Monocyte-Released Cytotoxic Factor: Effect on Various Cellular Functions, and Dependency of Cytolysis on Various Metabolic Processes
Authors: Nissen-Meyer Jon , Kildahl-Andersen Odd , Austgulen Rigmor
Published: January 2018

The ability of the human monocyte-released cytotoxic protein factor(s) (CF) to mediate cytolysis when various metabolic processes in target cells were inhibited, and its effect on various cell functions have been studied. Cytolysis was reduced when target cells were exposed to either chloroquine or colchicine, indicating that lysosome function and the formation of microtubule are of importance with respect to the interaction between CF and target cells, possibly by effecting internalization and processing CF and its receptor. A decrease in the energy charge of the target cells did not affect the ability of CF to mediate cytolysis, since the 60-70% reduction of the cellular ATP content by uncoupling oxidative phosphorylation had little effect on CF-induced lysis. It was unnecessary for target cells to be actively growing and dividing for CF to induce lysis because inhibition of cellular protein, RNA, and DNA synthesis potentiated the cytolytic effect of CF.

CF-induced cell lysis of Walter and Eliza Hall Institute (WEHI) 164 target cells was first detected between 3.0 and 4.5 hr after addition of CF. Target cell DNA synthesis nearly terminated within 3-4 hr, about the time of first cell lysis detection. Compared to the effect on DNA synthesis, the CF effect on protein synthesis was negligible and only a slight reduction was detected in the RNA synthesis and in the cellular ATP content. The results indicated that the DNA-synthesizing machinery may either be a primary target site of CF or a rapid influence of CF action on its primary target site; however, neither the protein- or RNA-synthesizing machinery nor the cellular ATP generating systems are likely to be primary target sites.
Prostaglandin E2 Content of Regressing and Progressing Moloney Sarcomas
Authors: Wechsler Sally J , Eurell Thomas E , Russell Stephen W
Published: January 2018

Regressing mouse Moloney sarcomas contain macrophages that are activated for tumor cell killing, while those found in progressively growing sarcomas either cannot kill or do so very poorly. Prostaglandin E2 (PGE2) has been shown to down-regulate macrophage activation in vitro. The study described here was designed, therefore, to ascertain and compare the concentrations of PGE2 in regressing and progressing Moloney sarcomas. Tumors were harvested for extraction and analyzed using conditions that minimized artifactual increases in PGE2 levels attributable to de novo synthesis. Concentrations of PGE2 were higher in progressing, compared to regressing, Moloney sarcomas during the early stages of tumor development. At nearly all time points, however, whether the neoplasms were of regressing or progressing type, the estimated concentrations of PGE2 in tumors exceeded the level that completely inhibits macrophage activation for tumor cell killing in vitro, that is, 10−8 M. These data suggest either that PGE2 is not responsible for down-regulating activation in Moloney sarcomas or that, if PGE2 is responsible for the negative regulation of activation in progressing Moloney sarcomas, there must be something in regressing sarcomas that prevents the hormone from having its inhibitory effect.
Phagocytic Rabbit Cell Lines Expressing Class II MHC Products: Establishment of Cell Lines by Viral Transformation
Authors: Kulaga Henrietta , Kindt Thomas J , Sogn John A
Published: January 2018

Continuous rabbit macrophage-like cell lines were established after in vitro infection of spleen cells with either Simian virus 40, lymphotropic papovavirus or herpesvirus sylvilagus. These cell lines are characterized as morphologically similar to mature macrophages, esterase positive, and have unrearranged immunoglobulin genes. They possess macrophage functionality because they are highly phagocytic and are able to mediate an antibody dependent cell mediated cytotoxicity assay on chicken red blood cells. Northern blot analysis of total cellular RNA with a DQα probe indicates that the cell lines constitutively express message for class II gene products. In addition, cell sorter analysis indicates that two of these lines display class II antigen at the cell surface.
Extracellular Calcium Is not an Absolute Requirement for Tumoricidal Activation of RAW-264 Macrophage-Like Cell Line
Authors: Gorecka-Tisera Alicja M , McCulloch Mary Ann
Published: January 2018

The purpose of these studies was to establish whether extracellular calcium (Cao  2+) plays a role in the process of activation of RAW-264 macrophages for tumor cell killing. We found that these cells were capable of developing a significant level of cytolytic activity under treatment with lymphokine (LK) and lipopolysaccharide (LPS), in the absence of Cao  2+ and that responses developed in Ca2+-free media were only 6–18% lower in comparison with the responses developed in the presence of Cao  2+. The determination of 45calcium uptake in RAW-264 cells treated with LK and LPS showed that the rate of 45calcium uptake has displayed no increase during either the course of activation or in activated, highly cytolytic cells. Finally, three calcium channel blockers examined here: verapamil, diltiazem and flunarizine, with concentrations ranging from 1 × 10-7 M – 2.5 × 10-5 M, showed no inhibitory effect on the process of activation. Nifedipine, another calcium channel blocker, inhibited the development of cytolytic activity with concentrations ranging from 1 × 10-6 M – 2.5 × 10-5 M. It could be argued, however, that this inhibition was nonspecific, since this agent was 13 times more potent with regard to the calcium ionophore A23187-induced release of β-glucuronidase, the function which is entirely dependent on Cao  2+. Taken together, these results suggest that Cao  2+ is not an absolute requirement for the process of tumoricidal activation of RAW-264 macrophages but it may play some supportive role in this process.
Effect of Bacterial Flora and Mouse Genotype (Euthymic or Athymic) on Scrapie Pathogenesis
Authors: Wade W F , Dees C , German T L , Marsh R F
Published: January 2018

Euthymic and athymic female BALB/c mice, reared under either germfree or defined flora conditions, were used to investigate the pathogenesis of scrapie after intracerebral or intraperitoneal inoculation. Time in days to onset of clinical signs (Stage I), to endstage (Stage II), and the time interval between Stage I and Stage II were compared among groups. In addition, scrapie agent titers in spleen were determined at 28 and 90 days after infection, as were agent titers in spleen and brain at Stage II. Three-way analysis of variance indicated that the bacterial flora, the presence or absence of a thymus, and the route of agent inoculation interact to produce significant differences in the pathogenesis of disease. The three factors in the experimental design also influenced the spleen titers of scrapie infectivity. The variation in scrapie pathogenesis among the groups of mice is likely to be mediated by differences in their reticuloendothelial systems. These differences may alter the agent's adsorption in spleen and/or route of transport from spleen to brain.
Size and Filterability of Human and Hamster Pulmonary Macrophages Exposed to Cigarette Smoke
Authors: Smith Clark M. , Tukey David P , Boyd Ruth , Garlich Dawn J , Hoidal John , Clawson C C
Published: January 2018

Pulmonary alveolar macrophages (PAM) obtained from healthy human cigarette smokers generated greater pressure in filtration through small cylindrical pores than PAM from nonsmoking controls. A well standardized hamster smoking model was employed to study the structural basis for the impaired PAM filtration acompanying cigarette smoke exposure. Hamsters also demonstrated increased PAM filtration pressure through cylindrical pores less than half a cell diameter in size after inhalation of cigarette smoke, but not after exposure to vapor phase smoke that had been strained through a particle filter. Cigarette smoker PAM were equally filterable as control through larger diameter channels, and showed comparable improvements in filterability with cytochalasin B treatment. Altered PAM filtration was associated with the formation of characteristic cytoplasmic smoker inclusions, an increase in cell size, and loss of redundant, ruffled surface membrane. The study of smoker and control PAM separated into different sizes on the basis of variations in cell density suggested that discrepancies in cell size were insufficient to explain the filtration disparity. A loss of availability of surface membrane for deformation appeared to be the most important factor responsible for the impaired filtration.
Immunohistochemical Analysis of the Involvement of F4/80 and Ia-Positive Macrophages in Mouse Liver Infected With Lymphocytic Choriomeningitis Virus
Authors: Dixon Jane E , Allan Jane E , Doherty Peter C , Hume David A
Published: January 2018

The distribution of cells bearing the F4/80 macrophage marker and class II MHC glycoproteins has been analyzed in the livers of mice infected with viscerotropic LCMV. The number of F4/80+ macrophages in the liver increases greatly during infection. Much of this is due to the localization of F4/80+ Ia+ monocytes, large numbers of which attach to the walls of the sinuses and the central and hepatic veins. Numerous lymphocytes are also observed in the sinusoids, frequently in close association with macrophages. The lymphocytes tend to move on from this intravascular location, while the macrophages remain. Foci of F4/80− Ia− mononuclear cells (probably T lymphocytes) are found both in the liver parenchyma and in locations that are obviously perivascular. The most prominent of these lymphocyte cuffs are located in the region of the portal triad. Infiltration of the lymphocyte foci with F4/80+ Ia+ elements occurs late, concurrent with evidence of cell death within the lesion. The dissociation in the focal accumulations of lymphocytes and monocyte/macrophages early in the disease process is both striking and unforeseen.
Enhanced Superoxide Production by Rat Alveolar Macrophages Stimulated In Vitro With Biological Response Modifiers
Authors: Badger Alison M
Published: January 2018

The kinetics of superoxide release and the effects of several biological response modifiers (BRM) on superoxide release from rat pulmonary alveolar macrophages (AM) have been studied. These cells produced superoxide anion both spontaneously and in repsonse to phorbol myristate acetate (PMA) in a dose-related manner. The response to PMA peaked in approximately 2 hr and maintained plateau levels for an additional 2-3 hr before subsiding. Pretreatment of the macrophages in vitro with a number of immunostimulants enhanced the production of superoxide above that of controls. The release of superoxide in response to the immunostimulants was a slow phenomenon that took place over a 3-5 hr time period. Lymphokine-containing supematants from concanavalin A (con A)-stimulated rat spleen cells (LK-Sup), murine recombinant gamma interferon (rMuIFN-γ), nigeran, and muramyl dipeptide (MDP) enhanced this response in a dose-related manner. Poly I:C and Salmonella typhosa lipopolysaccharide (LPS) stimulated rat alveolar macrophages at low but not high concentrations. In contrast to the alveolar macrophages, rat peritoneal exudate cells were not activated by immunostimulants to produce increased amounts of superoxide.
Enhancement of Human Peripheral Blood Monocyte Respiratory Burst Activity by Aggregated C-Reactive Protein
Authors: Zeller Janice M , Landay Alan L , Lint Thomas F , Gewurz Henry
Published: January 2018

We had previously demonstrated that C-reactive protein (CRP), an acute phase reactant, when aggregated or coupled to a ligand, interacts with monocytes and certain human peripheral blood lymphocytes. The purpose of the present study, after further characterizing the binding interaction of CRP with human monocytes, was to focus on the biological response of monocytes to CRP binding. Flow cytometric analysis of human mononuclear leukocytes, following incubation with fluoresceinated heat-aggregated CRP (Agg-CRP), revealed that while greater than 70% of monocytes bound Agg-CRP, only 8% of lymphocytes demonstrated positive fluorescence. Furthermore, mean channel fluorescence values indicated that monocytes bound greater amounts of Agg-CRP per cell than did lymphocytes. Luminol-enhanced chemiluminescence (CL) was used as a measure of monocyte respiratory burst activity. Monocytes were stimulated only minimally by Agg-CRP alone; however, Agg-CRP substantially enhanced the CL response to heat-aggregated IgG. This Agg-CRP enhancing effect was selective for IgG-initiated monocyte activation, as no augmentation in CL was observed following cell stimulation with phorbol myristate acetate or serum-opsonized zymosan. These results demonstrate that aggregated CRP binds to the major proportion of human monocytes and selectively augments Fc receptor-mediated stimulation of monocyte oxidative metabolism.
Natural Killer Cell Activity in the Rat. VI. Characterization of Rat Large Granular Lymphocytes as Effector Cells in Natural Killer and Antibody-Dependent Cellular Cytotoxic Activities
Authors: Fukui Hiroyasu , Overton W Roy , Herberman Ronald B , Reynolds Craig W
Published: January 2018

The present study examined rat natural killer (NK) celts, which mediate not only NK activity but also anitibody-dependent cellular cytotoxicity (ADCC). NK and ADCC activities were compared with regard to 1) organ distribution, 2) strain distribution, 3) Percoli fractionation of the effector cells, 4) effects of aging, and 5) potential to be augmented by biological response modifiers (BRM). Like NK activity, appreciable ADCC activity was observed in peripheral blood leukocytes (PBL), splenic leukocytes (SPL), and peritoneal exudate cells (PEC), but not in cell preparations from the peripheral lymph nodes (PLN), mesenteric lymph nodes (MLN), bone marrow (BM), and thymus (THY). ADCC activity, when compared with NK activity, was significantly higher in PBL but the same or lower in SPL and PEC. In terms of strain distribution, a high NK/AOCC strain (mu/mu), four intermediate NK and high ADCC strains (PVG/RTLRL, Lewis, PVG/OLA, and F344), an intermediate NK/ADCC strain (WF/N), and a low NK/ADCC strafa (Buffato) were observed.

Fractionation of effector cells on discontinuous Percoli gradients revealed that both NK and ADCC activities were associated with relatively high-density large granular lymphocytes (LGL). In contrast, ADCC but little or no NK activity was associated with tower density LGL. However, the NK activity of this tower-density LGL population could be elicited following the in vitro incubation with a number of BRM, including rat interferon (IFN) and OK-432, but not rat interteukin-2 (IL-2). In general, the ADCC activity of both higher and tower density LGL enriched celi populations correlated with both the frequency of FC7R+ LGL and the percentage of LGL binders to antibody-coated P815 target cells.

The present study also has shown that in contrast to NK activity, which remained relatively stable with age, ADCC activity from F344 but not WF/N rats increased until 30-50 wk of age. This increase of ADCC activity in older F344 rats was accompanied by an increase in the percentage and absolute number of lower density FCγR+ LGL.

This study demonstrates a number of similarities and differences between NK and ADCC activities in the rat. These findings should be useful for further examining and comparing the in vivo development and biological role of these two effector arms of the immune system.
1,25-Dihydroxyvitamin D3 Stimulates Interieukin-2 Production by a T Cell Lymphoma Line (MLA-144) Cultured in Vitamin D-Deficient Rat Serum
Authors: Haverty Thomas , Haddad John G , Neilson Eric G
Published: January 2018

A lymphocyte T celt line (MLA-144), which constitutively secretes interieulin-2 (IL-2), was shown to express receptors for 1,25-diydroxyvitamin D3 (1,25(OH)2 D3). The proliferation of an IL-2-dopondent cell line (HT-2) in response to supernatant! from MLA-144 cells was employed as an index of IL-2 production by MLA-144 cells. IL-2 production was two fold higher from MLA-144 cells cultured in 2% vitamin D-deficient rat serum compared to 10% fetal calf serum (FCS). The addition of 1,25(OH)2D3 at 10-15 Mor 10-11 M augmented IL-2 production by MLA-144 cells in vitamin D-deflcient rat serum, but not in fetal calf serum. At 10-7 M 1,25(OH)2D3 there was inhibition of IL-2 production by MLA-144 cells In either vitamin D-deflcient serum or FCS. There was no effect of 1,25(OH)2D3, added directly to HT-2 celis. Monoclonal antibody to the IL-2 receptor competitively inhibited the proliferation of HT-2 cells in response to MLA-144 supernatants, suggesting that it was IL-2 from the MLA-144 supernatants which influenced HT-2 proliferation. Our findings demonstrate biphasic does effects of 1,25(OH)2D3 on rymphokine secretion. The use of vitamin D deficient rat serum allowed us to demonstrate the effects of 1,25(OH)2D3 in the physiologic and subphysiologic range.
Metabolic Characterization of Mouse Bone Marrow Cells Responsive to Estrogenic Inhibition: Hexose Monophosphate Shunt Enzyme Activity in Enriched Populations of Mature Cells and Progenitor Cells
Authors: Dieter M P , French J E , Boorman G A , Luster M I
Published: January 2018

Compounds with estrogenic activity cause initial toxic responses in the bone marrow characterized by hypocellularity and stem cell myelotoxicity. To elucidate the biochemical nature of these toxic responses, bone marrow cells were collected from mice treated with pharmacological doses of estrogenic chemicals, separated into enriched cell populations, and the enzymatic responses of the individual cell types characterized. Female B6C3F1 mice were injected s.c. with five daily doses of 0.07-5.6 μmoles diethylstilbestrol (DES) or 17-β estradiol. At 4 days posttreatment body and organ weights were recorded and bone marrow was collected for enumeration and assay of stem cell proliferative responses and enzyme analyses. Treatment with higher dose levels of either estrogenic chemical caused equivalent thymic atrophy, but DES resulted in greater liver and spleen hypertrophy than estradiol. Hexose monophosphate shunt dehydrogenase enzymes in unfractionated bone marrow cells were more sensitive to inhibition by lower estrogen doses than representative enzymes from glycolysis of the Kreb's Cycle, and on an equimolar basis were inhibited to a greater extent by DES than by estradiol. Enzyme analyses after density gradient cell separation indicated that 70-80% of the hexose monophosphate shunt enzyme activity in bone marrow from untreated mice occurred in the enriched band of cells containing predominantly granulocyte-macrophages. The majority of the enzyme inhibition induced by DES treatment could also be ascribed to this cellular population. Furthermore, it was shown that DES had a greater inhibitory effect on the proliferative capacity of the committed stem cells than on the multipotential stem cell population, and the main response was again expressed in the enriched band of cells containing predominantly granulocyte-macrophage precursors. Preliminary endocrine ablation experiments indicated estrogen inhibition of hexose monophosphate shunt enzyme activity was independent of the adrenal and the ovary, but was mediated through the thymus at lower estrogen concentrations.
L-Fucose, D-Mannose, L-Galactose, and Their BSA Conjugates Stimulate Macrophage Migration
Authors: Takata I , Chida K , Gordon M R , Myrvik Q N , Ricardo M J , Kucera L S
Published: January 2018

The effect of selected monosaccharides on the random migration of normal adult rabbit alveolar macrophages (AM) was investigated. It was observed that 10 mM of L-fucose, L-galactose, or D-mannose stimulated AM migration 1.5-2.0 times. In addition, derivatives of L-fucose and D-mannose occupying the carbon-6 position such as L-fucosyl-lactose, D-mannose-6-phosphate, D-mannitol, and mannan enhanced the migration of AM, whereas derivatives of L-fucose and D-mannose in the carbon-1 position produced no migration enhancement. Macrophage migration enhancement activity that was produced spontaneously by spleen cell cultures from normal young rabbits was destroyed by treatment with L-fucosidase. Accordingly, the migration enhancement factor (MEF) found in spleen cell culture supernatants appeared to depend on L-fucose conjugated to some protein carrier because MEF was non-dialyzable. When normal adult AM were treated with L-fucosidase, they lost their responsiveness to migration inhibitory factor (MIF) but retained their responsiveness to MEF. We have interpreted this to mean that the MIF and MEF receptors are distinct. Synthetic MEFs were prepared by conjugating L-fucose, D-mannose, of L-galactose to bovine serum albumin (BSA). It was noted that these sugar-BSA conjugates were about 200 times more effective than the corresponding free sugars in producing migration enhancement. In addition, these sugar-BSA conjugates neutralized MIF activity in a migration inhibition test.
For when bacterial infections persist: Toll-like receptor-inducible direct antimicrobial pathways in macrophages
Authors: Stocks Claudia J , Schembri Mark A , Sweet Matthew J , Kapetanovic Ronan
Published: January 2018

Macrophages are linchpins of innate immunity, responding to invading microorganisms by initiating coordinated inflammatory and antimicrobial programs. Immediate antimicrobial responses, such as NADPH-dependent reactive oxygen species (ROS), are triggered upon phagocytic receptor engagement. Macrophages also detect and respond to microbial products through pattern recognition receptors (PRRs), such as TLRs. TLR signaling influences multiple biological processes including antigen presentation, cell survival, inflammation, and direct antimicrobial responses. The latter enables macrophages to combat infectious agents that persist within the intracellular environment. In this review, we summarize our current understanding of TLR-inducible direct antimicrobial responses that macrophages employ against bacterial pathogens, with a focus on emerging evidence linking TLR signaling to reprogramming of mitochondrial functions to enable the production of direct antimicrobial agents such as ROS and itaconic acid. In addition, we describe other TLR-inducible antimicrobial pathways, including autophagy/mitophagy, modulation of nutrient availability, metal ion toxicity, reactive nitrogen species, immune GTPases (immunity-related GTPases and guanylate-binding proteins), and antimicrobial peptides. We also describe examples of mechanisms of evasion of such pathways by professional intramacrophage pathogens, with a focus on Salmonella, Mycobacteria, and Listeria. An understanding of how TLR-inducible direct antimicrobial responses are regulated, as well as how bacterial pathogens subvert such pathways, may provide new opportunities for manipulating host defence to combat infectious diseases.
In vivo CD30 expression in human diseases with predominant activation of Th2-like T cells
Authors: D'Elios M M , Romagnani P , Scaletti C , Annunziato F , Manghetti M , Mavilia C , Parronchi P , Pupilli C , Pizzolo G , Maggi E , Del Prete G F , Romagnani S
Published: January 2018

CD3O is a member of the tumor necrosis factor (TNF) receptor family, originally described as a marker for Hodgkin and Reed-Sternberg cells in Hodgkin's disease, which has been found to be preferentially expressed by T cells producing Th2-type cytokines. The presence of CD3O expression was assessed by both immunohistochemistry and reverse transcriptase-polymerase chain reaction in the target organs of patients with Th1- or Th2-dominated disorders. CD3O expression was found in neither the gut of patients with Crohn's disease nor in the gastric antrum of Helicobacter pylori-infected patients, where there was high interferon-γ (IFN-γ) expression. In contrast, high CD3O expression in the apparent absence of IFN-γ expression was observed in the skin of patients with systemic sclerosis or chronic graft versus host disease (GVHD), which can be considered Th2-dominated disorders. Moreover, high levels of soluble CD3O were found in the serum of both systemic sclerosis and GVHD patients but not in the serum of patients suffering from multiple sclerosis, a Th1-dominated disorder. Thus, CD3O expression appears to be preferentially associated with Th2-type responses not only in vitro but also in vivo.
Wherefore interferon?
Authors: Gresser Ion
Published: January 2018

Interferon (IFN) was discovered by virologists interested in the phenomenon of viral interference and for many years was considered to be exclusively an antiviral substance. In time, it was accepted that the antiviral action was only one manifestation of the multiple effects of IFN on cells. IFN was shown to inhibit the development of tumors, to modulate immune function, and even to induce disease. Some aspects of these phenomena will be discussed. Despite this plethora of biological effects of type I IFN, the potency of its antiviral action, combined with its varied effects on both the cell-mediated and humoral immune response, render it a most effective and remarkable antiviral substance. Arguments will be presented to support the hypothesis that the major role of IFN is, after all, in antiviral defense. Finally, the use of type I IFN in the treatment of patients with viral and neoplastic diseases and even diseases of varied and unknown etiology is discussed.
Coupling of neutrophil apoptosis to recognition by macrophages: coordinated acceleration by protein synthesis inhibitors
Authors: Whyte Moira K B , Savill John , Meagher Laura C , Lee Alison , Haslett Christopher
Published: January 2018

Onset of apoptosis in many cell types, including the neutrophil granulocyte, leads to recognition and ingestion by macrophages, a key regulatory step in clearance of inflammatory cells from inflamed sites. These studies examined the requirement for protein synthesis in neutrophil apoptosis and in the recognition of apoptotic neutrophils by monocyte-derived macrophages. Treatment with cyclohexidmide or actinomycin D produced a time- and concentration-dependent acceleration of apoptosis in populations of neutrophils purified from human peripheral blood. Both compounds caused significant promotion of apoptosis after 8 h (apoptosis was 7.7 ± 2.9%, mean ± sem, in control populations, 57.5 ± 4.9% in cycloheximide-treated, and 73.4 ± 5.5% in actmomycin D-treated populations, n = 4, P < 0.001), which was associated with loss of neutrophil functional ability (assessed by shape change on N-formyl-methionyl-leucyl-phenylalanine stimulation) and increased macrophage recognition and ingestion of neutrophil populations with accelerated apoptosis. These results support the existence of survival proteins, which act as intracellular suppressors of programmed cell death. However, protein synthesis was not required for the recognition process because macrophage recognition was increased pari passu with the morphology of apoptosis.
5-Lipoxygenase inhibitors potentiate effects of TGF-β1 on the differentiation of human leukemia HL-60 cells
Authors: Kozubík Alois , Hofmanová Jirina , Dušek Ladislav , Musilová Eva
Published: January 2018

It was clearly demonstrated that two structurally different inhibitors of 5-lipoxygenase (5-LPO) metabolism (leukotriene synthesis), i.e. MK-886 and esculetin, when combined with transforming growth factor-β1 (TGF-β1), significantly enhanced the differentiation but did not change proliferation (i.e. cell number and cell cycle parameters) of human leukemia HL-60 cells in vitro. Although cell morphology and measurement of cell surface antigens (CD11b, CD14, and CD66b) after 48 h of combined treatment with MK-886 and TGF-β1 suggested a shift of the HL-60 cell population into more differentiated stages of myelopoiesis, cells with a fully mature phenotype were not observed. The effects on differentiation were better distinguished in the functional parameters of differentiation, i.e. oxidative burst of cells as detected by chemiluminescence and the nitroblue tetrazolium reduction test. Detailed statistical analysis of these data proved a significant synergistic interaction between TGF-β1 and inhibitors of 5-LPO metabolism. The differentiation effects of TGF-β1 alone and especially of its combination with MK-886 were most pronounced when the cells were pretreated with dimethyl sulfoxide or all-trans-retinoic acid. The results imply that either the lack of 5-LPO metabolites or a certain type of mis-balance in arachidonic acid metabolism can modulate the TGF-β1 effect on myeloid differentiation.
Differential regulation of biosynthesis of cell surface and secreted TNF-α in LPS-stimulated murine macrophages
Authors: Chaudhri Geeta
Published: January 2018

Activated macrophages synthesize a 26-kDa cell surface form and a 17-kDa secreted form of tumor necrosis factor α (TNF-α). This study was designed to investigate possible differences between the biosynthesis of these two forms by murine peritoneal exudate cells (PEC) and a murine macrophage cell line (RAW 264.7) stimulated with lipopolysaccharide (LPS). Both PEC and RAW 264.7 cells produced surface and secreted TNF-α in response to LPS in a dose-dependent manner. However, much lower concentrations of LPS (100 ng/mL) were needed for optimal expression of surface TNF-α than for secreted TNF-α (1 μg/mL). Furthermore, concentrations of actinomycin D that inhibit the synthesis of new mRNA and the production of secreted TNF-α did not block the expression of surface TNF-α on LPS-stimulated cells. Cycloheximide inhibited the production of both forms of TNF-α at similar concentrations. The effects, on the expression of the surface form of TNF-α, of various pharmacological agents known to inhibit the production of secreted TNF-α were studied. It was found that: (1) dexamethasone, a glucocorticoid agonist and (2) ETI and ETYA, inhibitors of lipoxygenase, had no effect on cell surface TNF-α at concentrations that inhibited secreted TNF-α. The data show that there are differences in the production of surface and secreted TNF-α and indicate that the regulation of synthesis of this protein is more complex than that suggested by a mere precursor/product relationship between the two forms.
Microorganisms and their interaction with the immune system
Authors: Kotwal Girish J
Published: January 2018

Microorganisms interact with the immune system in multiple ways. In an interaction between a microorganism and its host, the defense of the host does not go unchallenged. Microorganisms have for decades been suspected of possessing the capabilities of hiding from and escaping the consequences of immune surveillance. Escape mechanisms like antigenic variation, latency, and genomic integration can best be described as passive mechanisms for avoiding interaction with the host immune system, to differentiate them from the more engaging and host-directed active mechanisms of interaction. Studies of the mechanism of direct entry of viruses (HIV, measles, and enteroviruses), bacteria (streptococci and staphylococci), and parasites (Leishmania and plasmodium) into immune cells like CD4+ T cells or macrophages, as reported very recently, indicate an even more aggressive mode of interaction. This aggressive mechanism of interaction with the components of the host immune system allows the microbe not only to block the normal function of immune components on the surface of immune cells from functioning, but also to obliterate a vital immune function, cellular immunity, causing immunosuppression, e.g. the depletion of CD4+ T cells due to the entry and replication of HIV. Collectively, microorganisms have evolved various mechanisms by which they can actively block almost any cellular, humoral, or systemic immune response. One general feature of the proteins that assist microorganism to immunomodulate and actively evade host defense is their structural and therefore functional similarity to the host proteins, which they effectively mimic. Understanding the different mechanisms by which microorganisms interact with the immune system can impact the design of live vaccines as well as the development of novel therapeutic immunomodulators that can provide medicine with powerful new tools to manage immune disorders, allograft rejection, remote multiple organ failure resulting from trauma, autoimmune diseases, etc.
Humanized mAb H22 binds the human high affinity Fc receptor for IgG (FcγRI), blocks phagocytosis, and modulates receptor expression
Authors: Wallace Paul K , Keler Tibor , Coleman Kimberly , Fisher Jan , Vitale Laura , Graziano Robert F , Guyre Paul M , Fanger Michael W
Published: January 2018

About 10–15% of patients with immune thrombocytopenic purpura (ITP) cannot be controlled by corticosteroid therapy and splenectomy. For these patients treatment with high-dose IVIgG induces partial or complete responses. The clinical benefits of IVIgG could be due to blockade of Fc receptors for IgG (FcγR), because several model systems clearly show that functional FcγR are essential for establishment of ITP and related diseases. However, the specific contributions of the three individual classes of FcγR remain to be more completely defined. Recently monoclonal antibody (mAb) H22, which recognizes an epitope on FcγRI (CD64) outside the ligand binding domain, was humanized by grafting its complementarity determining regions onto human IgG1 constant domains. Because FcγRI has a high affinity for human IgG1 antibodies, we predicted mAb H22 would also bind to FcγRI through its Fc domain and block FcγRI-mediated phagocytosis. These studies demonstrate that mAb H22 blocked phagocytosis of opsonized red blood cells 1000 times more effectively than an irrelevant IgG. Moreover, cross-linking FcγRI with mAb H22 rapidly down-modulated FcγRI expression on monocytes without affecting other surface antigens. We conclude that because mAb H22 is a humanized mAb that blocks the FcγRI ligand binding domain and down-modulates FcγRI expression, it is a particularly good candidate for evaluating the role of FcγRI in patients with ITP.
In vitro expansion of CD13+CD33+ dendritic cell precursors from multipotent progenitors is regulated by a discrete fas-mediated apoptotic schedule
Authors: Santiago-Schwarz Frances , Borrero Michelle , Tucci John , Palaia Thomas , Carsons Steven E
Published: January 2018

We provide new information on how apoptosis regulates the expansion and survival of dendritic cell (DC) elements during in vitro hematopoiesis. Functionally distinct apoptotic schedules were associated with different phases of DC development when multipotent CD34+ progenitor cells were treated with GM-CSF + TNF ± SCF (c-kit ligand). During early phases of growth, unselected progenitors underwent apoptosis. During intermediate stages, high levels of apoptosis resulted in the preferential selection of DC precursors, as revealed by the massive expansion of DR+CD33+CD13+ cells. Late apoptosis was associated with the death of mature DCs. Apoptotic events surrounding the earlier periods were related to the exogenous addition of TNF-α and appeared to be mediated by fas. In contrast, those events associated with terminally differentiated DCs were fas independent because there was no correlation between fas expression and cell death. The bcl-2 protein family appeared to confer resistance to apoptotic death, as revealed by the high levels of bcl-2 and bclxL during peak DC development and in long-term DC cultures. We demonstrate that activation of distinct apoptotic programs regulates DC development and homeostasis. Although suppression of apoptosis may prolong the survival of late DC elements, an earlier apoptotic schedule appears to be required for the selective expansion of DC elements from multipotent progenitors. Our data also provides insight into the mechanism(s) of myeloid lineage selection by cytokines such as TNF-α, which may promote both cell death and survival.
Ultraviolet irradiation accelerates apoptosis in human polymorphonuclear leukocytes: protection by LPS and GM-CSF
Authors: Sweeney John F , Nguyen Phu Kim , Omann Geneva M , Hinshaw Daniel B
Published: January 2018

Polymorphonuclear leukocytes (PMN) play a central role in host response to injury and infection. Understanding factors that regulate PMN survival may therefore have a major influence on the development of novel treatment strategies for controlling life-threatening infections, as well as local and systemic inflammatory responses. Unfortunately, the presently utilized in vitro culture model of PMN apoptosis makes the examination of early biochemical events surrounding PMN apoptosis very difficult. This study demonstrates that a short course of UV irradiation (15 min) can be used to induce rapid progression of PMN through the apoptotic process with 70–90% of PMN displaying features of apoptosis by 4 h after UV exposure. Bacterial lipopolysaccharide and granulocyte-macrophage colony-stimulating factor, which are known to prolong PMN survival during in vitro culture, also protected PMN from UV-accelerated apoptosis. The UV-accelerated model of PMN apoptosis provides another valuable tool for the investigation of early signaling pathways associated with inducing or delaying PMN apoptosis.
Secreted poxvirus chemokine binding proteins
Authors: Lalani Alshad S , McFadden Grant
Published: January 2018

Poxviruses encode a variety of immunomodulatory proteins that subvert the cytokine networks of infected hosts. Myxoma virus, a poxvirus pathogen of rabbits, expresses two distinct 35- to 40-kDa secreted glycoproteins that bind a broad spectrum of chemokines. The first of these, designated M-T7, is encoded by the T7 gene and is the first example of what is here referred to as type-I chemokine binding protein (CBP-I). M-T7 was initially discovered as a secreted viral homologue of cellular interferon-γ receptor but binding studies indicate that purified M-T7 protein also interacts with members of the CXC, CC, and C chemokine families through the conserved heparin-binding domains. The second myxoma protein, M-T1, also called CBP-II, is a member of a larger superfamily of poxvirus proteins that includes related secreted 35-kDa proteins encoded by a wide variety of orthopoxviruses. Deletion analysis of either CBP-I or -II genes within recombinant poxvirus constructs revealed profound alterations in the trafficking of infiltrating leukocytes into virus-infected lesions. It is proposed that the interaction of CBP-I with the conserved heparin-binding domains found on most chemokines represents a novel mechanism for altering multiple chemokine functions in vivo. In summary, CBP-I and CBP-II are the first examples of secreted virus proteins that bind to multiple chemokine family members as part of a strategy to prevent the early phase of inflammatory cell migration into virus-infected tissues.
In vivo properties of monocyte chemoattractant protein-1
Authors: Gu Long , Rutledge Barbara , Fiorillo Joseph , Ernst Catherine , Grewal Iqbal , Flavell Richard , Gladue Ron , Rollins Barrett
Published: January 2018

Monocyte chemoattractant protein-1 (MCP-1) attracts monocytes, memory T lymphocytes, and natural killer (NK) cells in vitro. Its expression has been documented in disorders characterized by mononuclear cell infiltrates, suggesting that it may contribute to the inflammatory component of such diseases as atherosclerosis, multiple sclerosis, or rheumatoid arthritis. To prove a causal association, the in vivo properties of MCP-1 must be understood. Several lines of transgenic mice have been constructed to address this question. A transgenic line in which MCP-1 expression is controlled by the MMTV-LTR expressed high levels of MCP-1 in multiple organs but showed no evidence for monocyte infiltration. Instead, these mice were more susceptible to infection by the intracellular pathogens, Listeria monocytogenes and Mycobacterium tuberculosis. These mice had high serum levels of MCP-1, suggesting that their circulating monocytes may have been desensitized or that MCP-1 stimulated a Th2-dominant response. In contrast, another model in which MCP-1 expression was controlled by the insulin promoter demonstrated a monocytic infiltrate in pancreatic islets. These results indicate that MCP-1 expression at low levels in an anatomically confined area results in monocyte infiltration, suggesting that when properly expressed, MCP-1's in vitro properties are reproduced in vivo. This justifies the examination of MCP-1-deficient mice in disease models in order to explore MCP-1's role in pathogenesis.
Mechanism and biological significance of constitutive expression of MGSA/GRO chemokines in malignant melanoma tumor progression
Authors: Luan Jing , Shattuck-Brandt Rebecca , Haghnegahdar Hamid , Owen James D , Strieter Robert , Burdick Marie , Nirodi Chaitanya , Beauchamp Dan , Johnson Kimberly Newsom , Richmond Ann
Published: January 2018

By reverse transcriptase-polymerase chain reaction, enzymelinked immunosorbent assay, and immunohistochemistry, MGSA-α, -β, -γ, and CXCR2 mRNA expression and proteins are detected in 7 out of 10 human melanoma lesions. The biological consequence of constitutive expression of the MGSA/GRO chemokine in immortalized melanocytes was tested in SCID and nude mouse models. Continuous expression of MGSA/GRO-α, -β, or -γ in immortalized melan-a mouse melanocytes results in nearly 100% tumor formation for each of the clones tested, whereas clones expressing only the neomycin resistance vector form tumors <10% of the time. Moreover, antibodies to the MGSA/GRO proteins slow or inhibit the formation of tumors in the SCID mouse model and block the angiogenic response to conditioned medium from the tumor-producing clones. Transcription of the MGSA/ GRO chemokines is regulated by an enhancesomelike complex comprised of the nuclear factor-κB (NF-κB), HMG(I)Y, IUR, and Sp1 elements. In Hs294T melanoma cells the half life of the IκB protein is shortened in comparison to normal retinal epithelial cells, facilitating the endogenous nuclear localization of NF-κB. We propose that this endogenous nuclear NF-κB, working in concert with the 115-κDa IUR-binding factor, promotes constitutive expression of MGSA/GRO genes.
Chemokines in neurological disease models: correlation between chemokine expression patterns and inflammatory pathology
Authors: Ransohoff Richard M
Published: January 2018

We have recently determined chemokine expression profiles in a variety of models of neural trauma and immune-inflammation. The results indicate the following: (1) Chemokine expression in posttraumatic inflammation is generally restricted to the monocyte chemoattractant MCP-1, and occurs before hematogenous cell entry into neural tissues. Therefore MCP-1 is an excellent candidate for a mediator of leukocyte recruitment in these settings. (2) Chemokine expression in immune-inflammation is diverse and includes both α- and β-chemokines. Chemokine production can be attributed to parenchymal and infiltrating cell populations. Early signs of inflammation precede chemokine expression, which is believed to exert the crucial function of amplifying the immunemediated inflammatory reaction. These observations provide a basis for evaluating model neurological disorders in transgenic mice that express chemokines ectopically or in mice that are deficient in chemokine ligands or receptors as a consequence of gene targeting. Ultimately, a clear definition of roles of chemokines and their receptors in neurological diseases will suggest rational intervention. J.