PubMed Health⌕ Search

Biomedical subjects

R M Strieter

Publications and source records attributed to R M Strieter.

At least 145 records · Page 8Linked to original sources
PubMed1997

Cell-associated human retinal pigment epithelium interleukin-8 and monocyte chemotactic protein-1: immunochemical and in-situ hybridization analyses.

Human retinal pigment epithelial (RPE) cells secrete chemokines, interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1) in response to pro-inflammatory cytokines. In this study we (1) examined the efficiency of human RPE IL-8 and MCP-1 secretion, (2) determined the amount of neutrophil and monocyte chemotactic activity in human RPE cell conditioned media and cell extracts that is attributable to IL-8 and MCP-1, respectively, and (3) assessed the sensitivity of immunohistochemistry and in situ hybridization for detecting chemokine production by cytokine-stimulated human RPE cells. Conditioned media and extracts from human RPE cells stimulated with various physiologic concentrations of interleukin-1 beta (IL-1 beta) (0.2-20 ng ml-1), tumor necrosis factor (TNF-alpha) (0.2-20 ng ml-1) or interferon-gamma (IFN-gamma) (10-1000 U ml-1) were examined to compare secreted and cell associated levels of IL-8 and MCP-1 at various time points up to 24 hr. ELISA demonstrated that IL-8 and MCP-1 are both efficiently secreted by pro-inflammatory cytokine treated human RPE cells. Substantial dose- and time-dependent RPE secretion of IL-8 was observed following stimulation with IL-1 beta or TNF-alpha, but cell associated IL-8 was detectable only after high dose (20 ng ml-1) IL-1 beta stimulation and comprised less than 1% of the total IL-8 induced. Dose- and time-dependent RPE cell MCP-1 secretion was also observed following IL-1 beta > TNF-alpha > IFN-gamma stimulation, with an average of 4% of the total MCP-1 retained within RPE. Bioassays demonstrated neutrophil and monocyte chemotactic activity in conditioned media from stimulated RPE cells, but not in human RPE cell extracts. Inhibition of conditioned media-induced chemotaxis by specific anti-IL-8 or anti-MCP-1 antibodies demonstrated that IL-8 and MCP-1 were responsible for the majority of HRPE-derived neutrophil (> 60%) and monocyte (53-57%) chemotactic activity, respectively. Using in situ hybridization IL-8 mRNA was readily detected within IL-1 beta > TNF-alpha stimulated RPE cells and MCP-1 mRNA easily visualized within IL-1 beta > TNF-alpha > or IFN-gamma stimulated cells. Immunohistochemistry to detect IL-8 was positive only in RPE cells exposed to high dose IL-1 beta (20 ng ml-1) for 8 or 24 hr and was weak. Immunohistochemical staining for MCP-1 in RPE cells was more intense and was visualized within RPE cells stimulated with IL-beta, TNF-alpha, or IFN-gamma. This study demonstrates that: (1) RPE cells efficiently secrete IL-8 and MCP-1 upon stimulation with pro-inflammatory cytokines; (2) secreted IL-8 and MCP-1 account for the majority of human RPE neutrophil and monocyte chemotactic activity; (3) in situ hybridization readily detects IL-8 and MCP-1 mRNA in cytokine stimulated RPE cells; and (4) immunohistochemistry demonstrates cell-associated MCP-1 in cytokine stimulated RPE cells, but only minimal cell-associated IL-8.

Cells, Cultured↗
PubMed1997

The C-X-C chemokine ENA-78 is preferentially expressed in intestinal epithelium in inflammatory bowel disease.

BACKGROUND & AIMS: Secretion of chemokines by epithelial cells may represent a crucial event in the pathogenesis of inflammatory bowel disease (IBD). Expression of the chemokine epithelial neutrophil-activating peptide 78 (ENA-78) was monitored in patients with IBD and normal controls. METHODS: In situ hybridizations were performed on 41 tissue specimens from 15 patients with IBD and 10 controls to detect ENA-78 messenger RNA (mRNA). Immunofluorescence stainings were used to localize ENA-78 protein. RESULTS: Intestinal epithelial cells expressing ENA-78 mRNA at detectable levels are found at comparable frequencies in patients with Crohn's disease and ulcerative colitis. Tissue specimens with mild to moderate histological signs of disease activity show slightly higher frequencies of ENA-78 mRNA-expressing epithelial cells than areas with signs of severe disease activity (P = 0.14). Immunofluorescence stainings showed presence of the ENA-78 protein in > 90% of preserved epithelial cells in IBD, in control tissues, ENA-78 mRNA was not detectable, and ENA-78 protein was detectable in 0%-30% of epithelial cells. CONCLUSIONS: The observations are in agreement with a role of the C-X-C chemokine ENA-78 in the pathogenesis of IBD.

Acute Disease↗
PubMed1997

Anti-P-selectin antibody decreases inflammation and thrombus formation in venous thrombosis.

PURPOSE: Venous thrombosis and inflammation are interrelated. P-selectin contributes to activation of leukocyte-mediated inflammation. Therefore, we hypothesized that the neutralization of P-selectin would decrease vein wall inflammation and thrombosis. METHODS: Twelve baboons underwent infrarenal inferior vena caval balloon occlusion to induce thrombosis. Two groups of four baboons received neutralizing intravenous anti-P-selectin antibody (PSab) GA6 or CY1748 before occlusion and at days 2 and 4. Four baboons received saline control injections. One baboon per group was killed at days 2, 6, and 13, and at 2 months. Analysis included phlebography, ultrasound, gadolinium (Gd)-enhanced magnetic resonance venography (reflecting vein wall inflammation), and histologic, morphometric, and protein evaluation of the vein wall. Thrombus presence or absence was assessed. RESULTS: By day 2 in PSab baboons, vein wall Gd enhancement was decreased in the mid-inferior vena cava and the right iliac vein (p < 0.05; GA6 vs control baboons), normalizing by 2 months. The mid-inferior vena cava revealed fewer neutrophils and total leukocytes in PSab baboons; however, for GA6 in the right iliac vein these decreases were not present despite the absence of Gd enhancement; they were decreased with CY1748. PSab baboons demonstrated significantly less thrombus than control baboons (p < 0.01, GA6 and CY1748 vs control baboons). CONCLUSIONS: Anti-P-selectin antibody decreases vein wall inflammation and thrombus formation. Inhibition of P-selectin may be useful in venous thrombosis prophylaxis.

Acute Disease↗
PubMed1997

Pathogenesis of venous thrombosis: a new insight.

Venous thrombosis and thrombophlebitis have long been observed to result in painful inflammation around the affected veins. The full extent of the synergistic interaction between thrombosis and the inflammatory response and how this leads to the later sequelae of chronic venous insufficiency is only now beginning to be understood. Venous thrombosis is known directly to elicit an inflammatory response in the thrombus and vein wall. Leukocytes including neutrophils and monocytes roll, adhere, activate and extravasate into the vein wall based on a vein wall cytokine/chemokine gradient producing an inflammatory response. Such a response leads to amplification of thrombus formation through mechanisms such as the elaboration of tissue factor on the surface of monocytes and the release of cathespin G from activated neutrophils (distrupting the endothelial cell barrier), exposing the thrombogenic subendothelial vein wall collagen. Selectins such as P-selectin and the proinflammatory cytokine tumor necrosis factor appear important in this vein wall response. Inhibition of inflammation before the initiation of the thrombotic event may decrease the detrimental vein wall changes that contribute to vein wall and vein valve damage and thrombus formation.

Chemokines↗
PubMed1997

Selective IL-10 inhibition of HLA-DR expression in IFN-gamma-stimulated human retinal pigment epithelial cells.

PURPOSE: Human RPE cells express HLA-DR antigens, bind leukocytes via ICAM-1, and secrete IL-8 and MCP-1, which attract and activate leukocytes. Since little is known concerning endogenous cytokines that may alter ocular immunologic and inflammatory mechanisms, we investigated whether IL-10, an immunosuppressive cytokine, modulates these HRPE features. METHODS: IL-10 effects on HLA-DR and ICAM-1 were examined by HRPE exposures to: (1) IFN-gamma (10-1000 U/ml) + IL-10 (1-100 U/ml) and (2) IL-10 pre-incubation followed by IFN-gamma + IL-10. Immunohistochemistry for HLA-DR and ICAM-1 was graded by masked observers. Flow cytometric analysis quantitated HRPE HLA-DR and ICAM-1. Effects of IL-10 on IL-1 beta (0.2 or 2 ng/ml)-, or TNF-alpha (0.2 or 2 ng/ml)-induced IL-8 and MCP-1 secretion and gene expression were assessed using enzyme-linked immunosorbent assay (ELISA) and Northern blot analysis. RESULTS: HLA-DR expression, detected by immunohistochemistry and flow cytometric analysis, showed dose-dependent increases to IFN-gamma. IL-10 pre-/co-incubation, but not co-incubation alone, markedly reduced HLA-DR expression, but did not modulate constitutive or IFN-gamma-induced ICAM-1. IL-10 alone did not induce MCP-1 or IL-8 secretion or steady-state mRNA expression, nor modulate IL-1 beta-, TNF-alpha- or IFN-gamma-induced IL-8 or MCP-1. CONCLUSIONS: This study suggests that HRPE HLA-DR antigens are selectively inhibited by IL-10, but the timing of IL-10 exposure may be crucial.

Chemokine CCL2↗
PubMed1997

In vitro transendothelial migration of blood T lymphocytes from HIV-infected individuals.

OBJECTIVE: We hypothesized that differential extravasation of circulating CD4+ or CD8+ T lymphocytes contributes to HIV-associated CD8+ lymphocytic alveolitis. Differences in T-cell transendothelial migration may be intrinsic or emerge at sites where vascular endothelium is activated by overexpression of tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma. DESIGN: We used an in vitro model of lymphocyte extravasation to assess transendothelial migration of peripheral blood mononuclear cells (PBMC) from HIV-positive individuals. We assayed bronchoalveolar lavage (BAL) fluid from HIV-positive and normal individuals to determine if increased levels of TNF-alpha and IFN-gamma were present in the lungs of HIV-infected individuals. METHODS: Transendothelial migration was assessed by determining the number and flow cytometric phenotype of PBMC adherent to or migrating across unstimulated or TNF-alpha and IFN-gamma-activated endothelial cell monolayers. We measured BAL fluid cytokine concentrations using standard antigen-capture enzyme-linked immunosorbent assays for TNF-alpha and IFN-gamma. RESULTS: T cells migrating across unactivated endothelial cells were significantly enriched for CD4+ T cells. Cytokine activation of endothelial cells allowed significantly greater transendothelial migration of CD8+ T cells compared to unactivated endothelial cells. TNF-alpha was increased in BAL fluid from HIV-positive individuals relative to controls. CONCLUSIONS: These data suggest that, in HIV-positive individuals, CD4+ T cells are migration competent and blood CD8+ T cells do not have enhanced migration competence relative to CD4+ T cells. CD8+ T cell extravasation is aided by TNF-alpha and IFN-gamma-induced endothelial cells activation.

Bronchoalveolar Lavage↗
PubMed1997

Anti-interleukin-12 therapy protects mice in lethal endotoxemia but impairs bacterial clearance in murine Escherichia coli peritoneal sepsis.

The overzealous production of proinflammatory cytokines in sepsis can result in shock, multiorgan dysfunction, and even death. In this study we assessed the role of endogenously produced interleukin (IL)-12 in murine models of endotoxemia and Gram-negative peritoneal sepsis. Initial studies indicated that intraperitoneal lipopolysaccharide (LPS) administration to mice induced a significant time-dependent increase in plasma, lung, and liver IL-12 levels. Passive immunization with anti-IL-12 serum intraperitoneally before LPS resulted in a marked reduction in plasma levels of tumor necrosis factor and interferon-gamma. Furthermore, we observed an increase in endotoxin-induced mortality in mice transiently overexpressing murine IL-12 using a recombinant adenoviral vector (Ad5 mIL-12) administered intraperitoneally. Neutralization of tumor necrosis factor or interferon-gamma in animals overexpressing IL-12 resulted in significant reductions in LPS-induced mortality, suggesting that the mechanism whereby IL-12 increases LPS-induced mortality is primarily mediated by the enhancement of these cytokines. In contrast, we observed no survival benefit in animals passively immunized with anti-IL-12 serum before the intraperitoneal administration of 2 x 10(8) live Escherichia coli. Interestingly, there was an approximately 70-fold increase in peritoneal fluid E. coli colony-forming units and the early onset of bacteremia in animals treated with anti-IL-12 serum, as compared with control animals. These results indicate that IL-12 is produced in response to LPS exposure, and the neutralization of this cytokine improves survival in endotoxin-challenged animals. However, IL-12 represents an essential component of antibacterial host defense, as anti-IL-12 therapy results in significant impairment in the host's ability to clear Gram-negative bacterial infection.

Animals↗
PubMed1997

Induction of gamma interferon production in human alveolar macrophages by Mycobacterium tuberculosis.

Gamma interferon (IFN-gamma) is a cytokine which plays a critical role in resistance to Mycobacterium tuberculosis infection. While T lymphocytes and natural killer cells are a major source of IFN-gamma, previous demonstrations that it can be produced by murine macrophages prompted us to examine the capacity of human alveolar macrophages to express IFN-gamma. Here we report that in vitro infection of alveolar macrophages with M. tuberculosis induces both the release of IFN-gamma protein and a transient increase in IFN-gamma mRNA levels. The IFN-producing cells were shown to be macrophages by reverse transcription-in situ PCR. We also observed that M. tuberculosis stimulation resulted in IFN-gamma-dependent expression of the chemokines IFN-gamma-inducible protein 10 and monokine induced by IFN-gamma, suggesting that macrophage-derived IFN-gamma can function in an autocrine and/or paracrine manner. The existence of a positive regulatory loop was suggested by the observation that exogenous IFN-gamma protein could induce IFN-gamma mRNA expression in uninfected alveolar macrophages. Interleukin-12 was also found to be a potent inducer of IFN-gamma production, and M. tuberculosis-induced IFN-gamma production appears to be mediated, at least in part, by IL-12. In contrast, M. tuberculosis-induced IFN-gamma production by alveolar macrophages could be blocked by exogenous interleukin-10. These studies are the first to demonstrate an autoregulatory role for IFN-gamma produced by alveolar macrophages infected in vitro with M. tuberculosis.

Cells, Cultured↗
PubMed1997

Alveolar macrophages are required for protective pulmonary defenses in murine Klebsiella pneumonia: elimination of alveolar macrophages increases neutrophil recruitment but decreases bacterial clearance and survival.

To study the in vivo role of alveolar macrophages (AM) in gram-negative bacterial pneumonia in mice, AM were eliminated by the intratracheal (i.t.) administration of dichloromethylene diphosphonate encapsulated liposomes. Subsequently, the AM-depleted mice were infected i.t. with 100 CFU of Klebsiella pneumoniae, and the effects of AM depletion on survival, bacterial clearance, and neutrophil (polymorphonuclear leukocyte [PMN]) recruitment were assessed. It was shown that depletion of AM decreases survival dramatically, with 100% lethality at day 3 postinfection, versus 100% long-term survival in the control group. This increased mortality was accompanied by 20- to 27- and 3- to 10-fold increases in the number of K. pneumoniae CFU in lung and plasma, respectively, compared to those in nondepleted animals. This decreased bacterial clearance was not due to an impaired PMN recruitment; on the contrary, the K. pneumoniae-induced PMN recruitment in AM-depleted lungs was sevenfold greater 48 h postinfection than that in control infected lungs. Together with an increased PMN infiltration, 3- and 10-fold increases in lung homogenate tumor necrosis factor alpha (TNF-alpha) and macrophage inflammatory protein 2 (MIP-2) levels, respectively, were measured. Neutralization of TNF-alpha or MIP-2, 2 h before infection, reduced the numbers of infiltrating PMN by 41.6 and 64.2%, respectively, indicating that these cytokines mediate PMN influx in infected lungs, rather then just being produced by the recruited PMN themselves. Our studies demonstrate, for the first time, the relative importance of the AM in the containment and clearance of bacteria in the setting of Klebsiella pneumonia.

Animals↗
PubMed1997

Nitric oxide is required for effective innate immunity against Klebsiella pneumoniae.

Nitric oxide (NO) has been associated with protection against various parasitic and viral infections and may play a similar role in bacterial infections. We studied the role of NO in host defense against Klebsiella pneumoniae infection in the lung. Initial studies demonstrated a time-dependent increase in NO production of the lungs of CBA/J mice following the intratracheal administration of K. pneumoniae (7 x 10(2) CFU). To assess the role of NO in Klebsiella pneumonia, mice were treated intraperitoneally with either L-NAME (N-omega-nitro-L-arginine methylester), a competitive inhibitor of NO synthesis, or D-NAME, an inert enantiomer. The treatment of Klebsiella-infected mice with L-NAME resulted in a 10- and 46-fold increase in K. pneumoniae CFU in lungs and blood, respectively, at 48 h post-K. pneumoniae inoculation compared to treatment of mice with D-NAME. In addition, a greater-than-twofold increase in mortality was evident in L-NAME-treated mice compared to the mortality in control animals. No significant difference in bronchoalveolar lavage inflammatory cell profiles was noted between L-NAME- and D-NAME-treated mice with Klebsiella pneumonia. Interestingly, increased levels of tumor necrosis factor, gamma interferon, macrophage inflammatory protein 1alpha (MIP-1alpha), and MIP-2 mRNA and protein were noted in infected mice treated with L-NAME compared to the levels in mice treated with D-NAME. Importantly, the in vitro incubation of murine alveolar macrophages with L-NAME, but not with D-NAME, resulted in a significant impairment in both the phagocytosis and killing of K. pneumoniae. In total, these results suggest that NO plays a critical role in antibacterial host defense against K. pneumoniae, in part by regulating macrophage phagocytic and microbicidal activity.

Animals↗
PubMed1997

Elevated levels of macrophage inflammatory protein 2 in severe murine peritonitis increase neutrophil recruitment and mortality.

We hypothesized that chemokines may play important roles in a cecal ligation and puncture (CLP) model of septic peritonitis in CD-1 mice. Concentrations of C-X-C (macrophage inflammatory protein 2 [MIP-2] and ENA-78) and C-C (MIP-1alpha and JE) chemokines were measured (by enzyme-linked immunosorbent assay) in serum, peritoneal lavage fluid, lung, and liver at 4, 8, 24, 48, and 96 h after CLP. Significant elevations in all measured chemokines occurred in peritoneal fluid after CLP (P < 0.05). MIP-2, in particular, increased dramatically (>400-fold, P < 0.001) in peritoneal fluid, serum, and to a lesser extent lung and liver (P < 0.05). Increased MIP-2 was correlated with severity of sepsis (P < 0.001). To determine the significance of this finding, mice were passively immunized prior to CLP with polyclonal antibody to MIP-2, which decreased mortality from 85 to 38% at 96 h (P < 0.01). To further understand the mechanism of the effect of MIP-2, additional measurements demonstrated that anti-MIP-2 prior to CLP decreased the percent neutrophils in peritoneal fluid (55% +/- 12%, compared with 82% +/- 10% in controls), but no significant changes in tumor necrosis factor alpha, interleukin-6, or interleukin-10 occurred. MIP-2 contributes to the inflammatory response and overall mortality in this model of severe septic peritonitis, possibly by increasing recruitment of neutrophils, which clear bacteria but may also injure the host.

Animals↗
PubMed1997

RANTES inhibits HIV-1 replication in human peripheral blood monocytes and alveolar macrophages.

Infection with human immunodeficiency virus (HIV)-1 most often leads to the development of acquired immune deficiency syndrome, which may manifest with opportunistic infections, many of which occur in the lung. Mononuclear phagocytes infected by HIV-1, being relatively resistant to its cytopathic effects, potentially act as a reservoir for the virus. The alveolar macrophage (AM), a differentiated lung tissue macrophage, is readily infected by HIV-1, after which the virus becomes relatively dormant. C-C chemokines, secreted by CD8 T lymphocytes and other cells, are known to suppress HIV replication in lymphocytes. In view of this observation, and the relative increase in CD8+ T lymphocytes during HIV-1 disease, particularly in the lung, we hypothesized that C-C chemokines might play a key role in suppressing HIV-1 replication in AM. We examined the effect of the C-C chemokines macrophage inflammatory protein (MIP)-1 alpha, MIP-1 beta, and regulated on activation normal T cell expressed and secreted (RANTES) singly and in combination on HIV-1 replication in peripheral blood monocytes (PBM) and AM infected in vitro. Our findings indicate that RANTES suppresses HIV-1 replication, as measured by reverse transcriptase activity, in PBM (41.3 +/- 15.2% of control, n = 3, P < 0.05) and AM (30.3 +/- 7.8% of control, n = 3, P < 0.05) in a dose-dependent manner. The other C-C chemokines had no significant effect singly (MIP-1 alpha PBM: 64.8 +/- 21.9%; AM: 115.0 +/- 2.4% of control; MIP-1 beta PBM: 68 +/- 19.6; AM: 63.3 +/- 26.2% of control) but modestly decreased HIV replication when incubated in addition to RANTES (24.5 +/- 6.5% of control). These observations suggest that RANTES plays a key role in modulating HIV-1 replication in mononuclear phagocytes in the blood and lung, and this may have therapeutic implications for prevention and/or treatment of HIV disease.

Chemokine CCL5↗
PubMed1997

Synchronous synthesis of alpha- and beta-chemokines by cells of diverse lineage in the central nervous system of mice with relapses of chronic experimental autoimmune encephalomyelitis.

Chemokines are secreted peptides that exhibit selective chemoattractant properties for target leukocytes. Two subfamilies, alpha- and beta-chemokines, have been described, based on structural, genetic, and functional considerations. In acute experimental autoimmune encephalomyelitis (EAE), chemokines are up-regulated systemically and in central nervous system (CNS) tissues at disease onset. Functional significance of this expression was supported by other studies; intervention with an antichemokine antibody abrogated passive transfer of EAE, and chemokines expressed in brains of transgenic mice recruited appropriate leukocyte populations into the CNS compartment. Chemokine expression in the more relevant circumstance of chronic EAE has not been addressed. We monitored the time course and cellular sources of chemokines (monocyte chemoattractant protein-1, macrophage inflammatory protein-1 alpha, interferon-gamma-inducible protein of 10 kd, KC, and regulated on activation, normal T-cell expressed and secreted cytokine) in CNS and peripheral tissues during spontaneous relapses of chronic EAE. We found coordinate chemokine up-regulation in brain and spinal cord during clinical relapse, with expression confined to CNS tissues. Monocyte chemoattractant protein-1, interferon-gamma-inducible protein of 10 kd, and KC were synthesized by astrocytic cells, whereas macrophage inflammatory protein-1 alpha and regulated on activation, normal T-cell expressed and secreted cytokine were elaborated by infiltrating leukocytes. The results demonstrate stringent regulation of multiple chemokines in vivo during a complex organ-specific autoimmune disease. We propose that chemokine expression links T-cell antigen recognition and activation to subsequent CNS inflammatory pathology in chronic relapsing EAE.

Animals↗