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Jonathan S Reichner

Publications and source records attributed to Jonathan S Reichner.

11 recordsLinked to original sources
PubMed2006

Beta-glucan is a fungal determinant for adhesion-dependent human neutrophil functions.

Candida albicans is a common cause of nosocomial infections whose virulence depends on the reversible switch from blastoconidia to hyphal forms. Neutrophils (or polymorphonuclear leukocytes (PMNs)) readily clear blastoconidia by phagocytosis, but filaments are too long to be ingested. Mechanisms regulating immune recognition and response to filamentous fungal pathogens are not well understood, although known risk factors for developing life-threatening infections are neutropenia or defects in the NADPH oxidase system. We show human PMNs generate a respiratory burst response to unopsonized hyphae. Ab specific for beta-glucan, a major component of yeast cell walls, blocks this response, establishing beta-glucan as a key molecular pattern recognized by PMNs in response to C. albicans. This study also elucidates recognition and signaling mechanisms used by PMNs in response to beta-glucan under conditions where phagocytosis cannot occur. Human PMNs adhered to immobilized beta-glucan and released an efficient plasma membrane respiratory burst. Ab blockade of the integrin complement receptor 3 (CD11b/CD18) significantly inhibited both of these functions. Furthermore, we show a role for p38 MAPK and actin but not protein kinase C zeta in generating the respiratory burst to beta-glucan. Taken together, results show that beta-glucan in C. albicans hyphae is accessible to PMNs and sufficient to support an innate immune response.

Actins↗
PubMed2006

The effect of PGG-beta-glucan on neutrophil chemotaxis in vivo.

The beta-glucans are long-chain polymers of glucose in beta-(1,3)(1,6) linkages, which comprise the fungal cell wall and stimulate cells of the innate immune system. Previous in vitro studies have shown the ability of beta-glucan to increase the chemotactic capacity of human neutrophils. The current study examined an in vivo correlate of that observation by testing the hypothesis that systemic beta-glucan treatment would result in enhanced migration of neutrophils into a site of inflammation and improve antimicrobial function. A model of acute inflammation was used in which polyvinyl alcohol sponges were implanted subcutaneously into the dorsum of rats. Animals treated with beta-glucan showed a 66 +/- 6% and 186 +/- 42% increase in wound cell number recovered 6 and 18 h postwounding, respectively. Increased migration did not correlate with increased chemoattractant content of wound fluid, alterations in neutrophil-induced loss of endothelial barrier function, or changes in neutrophil adhesion to endothelial cells. Systemic administration of SB203580 abrogated the enhanced migration by beta-glucan without altering normal cellular entry into the wound. Studies also showed a priming effect for chemotaxis and respiratory burst in circulating neutrophils isolated from beta-glucan-treated animals. Heightened neutrophil function took place without cytokine elicitation. Furthermore, beta-glucan treatment resulted in a 169 +/- 28% increase in neutrophil number and a 60 +/- 9% decrease in bacterial load in the bronchoalveolar lavage fluid of Escherichia coli pneumonic animals. Taken together, these findings demonstrate that beta-glucan directly affects the chemotactic capacity of circulating neutrophils through a p38 mitogen-activated protein kinase-dependent mechanism and potentiates antimicrobial host defense.

Animals↗
PubMed2005

Modulation of macrophage phenotype by soluble product(s) released from neutrophils.

The regulation of macrophage phenotype by neutrophils was studied in the s.c. polyvinyl alcohol sponge wound model in mice made neutropenic by anti-Gr-1 Ab, as well as in cell culture. Wounds in neutropenic mice contained 100-fold fewer neutrophils than those in nonneutropenic controls 1 day after sponge implantation. Wound fluids from neutropenic mice contained 68% more TNF-alpha, 168% more IL-6, and 61% less TGF-beta1 than those from controls. Wound fluid IL-10 was not different between the two groups, and IL-4 was not detected. Intracellular TNF-alpha staining was greater in cells isolated from neutropenic wounds than in those from control wounds. The hypothesis that wound neutrophil products modulate macrophage phenotype was tested in Transwell cocultures of LPS-stimulated J774A.1 macrophages and day 1 wound cells (84% neutrophils/15% macrophages). Overnight cocultures accumulated 60% less TNF-alpha and IL-6 than cultures of J774A.1 alone. The suppression of cytokine release was mediated by a soluble factor(s), because culture supernatants from wound cells inhibited TNF-alpha and IL-6 release from LPS-stimulated J774A.1 cells. Culture supernatants from purified wound neutrophils equally suppressed TNF-alpha release from LPS-stimulated J774A.1 cells. Wound cell supernatants also suppressed TNF-alpha and superoxide release from murine peritoneal macrophages. The TNF-alpha inhibitory factor has a molecular mass <3000 Da and is neither PGE2 nor adenosine. The present findings confirm a role for neutrophils in the regulation of innate immune responses through modulation of macrophage phenotype.

Animals↗
PubMed2005

Macrophage arginase regulation by CCAAT/enhancer-binding protein beta.

Arginase activity is expressed by macrophages in healing wounds and other sites of inflammation and has been shown to modulate the synthesis of nitric oxide, polyamines, and collagen. The role of CCAAT/enhancer-binding protein beta (C/EBPbeta) in the regulation of macrophage arginase by different agonists was investigated using C/EBPbeta-/- and +/+ macrophage cell lines. 8-Bromo-cyclic adenosine monophosphate (8-Br-cAMP, 0.5 mM), recombinant murine interleukin 4 (rmIL-4, 20 U/mL), Escherichia coli lipopolysaccharide (100 ng/mL), and hypoxia (1% O2) induced arginase activity in C/EBPbeta+/+ macrophages, where enzyme activity correlated with arginase I protein. Only rmIL-4 increased arginase activity in C/EBPbeta-/- cells. Arginase II protein was expressed constitutively in wild-type and C/EBPbeta-/- cell lines and was unaltered by 8-Br-cAMP or rmIL-4. rmIL-4-stimulated immortalized C/EBPbeta-/- macrophages demonstrated higher nuclear signal transducer and activator of transcription-6 (STAT6) and phospho-STAT6 content than their +/+ counterparts. Validating the biological relevance of findings with the cell lines, additional experiments examined wound fluids and peritoneal macrophages from C/EBPbeta-/- mice and demonstrated that both contained less arginase activity than those from wild-type controls. Wounds in C/EBPbeta-/- animals showed signs of delayed maturation, as manifested by the persistence of neutrophils in the inflammatory infiltrate. Peritoneal macrophages from C/EBPbeta+/+ animals responded to 8-Br-cAMP and rmIL-4 with increased arginase activity, whereas those from C/EBPbeta-/- mice did not respond to cAMP. Results demonstrate a key mechanistic role for C/EBPbeta in the modulation of macrophage arginase I expression in vivo and in vitro.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed2004

Transcriptional regulation of TNF-alpha production in neutropenia.

Neutropenia has been shown to markedly increase plasma TNF-alpha concentration after LPS injection and to enhance LPS-induced mortality. Experiments reported here demonstrate that the 15-fold higher plasma TNF-alpha concentration elicited by LPS in neutropenic vs. nonneutropenic unanesthetized mice correlated with increased hepatic and splenic, but not pulmonary, TNF-alpha mRNA. Core 2 beta-1,6-N-acetylglucosaminyltransferase-null and CD18-deficient mice also exhibited exaggerated plasma TNF-alpha responses to LPS injection. Findings suggest that extravasated neutrophils inhibit systemic TNF-alpha production and that they do so through organ-selective mechanisms involving CD18 integrin and selectin binding.

Animals↗
PubMed2004

The lectin-like domain of complement receptor 3 protects endothelial barrier function from activated neutrophils.

The adhesion of neutrophils to endothelial cells is a central event leading to diapedesis and involves the binding of the I-domain of beta(2) integrins (CD11/CD18) to endothelial ICAMs. In addition to the I-domain, the beta(2) integrin complement receptor 3 (CR3) (CD11b/CD18) contains a lectin-like domain (LLD) that can alter leukocyte functions such as chemotaxis and cytotoxicity. The present study demonstrates that, in contrast to the CR3 I-domain, Ab blockade of the CR3 LLD has no role in mediating neutrophil-induced loss of endothelial barrier function. However, activation of CR3 with the LLD agonist beta-glucan protects the barrier function of endothelial cells in the presence of activated neutrophils and reduces transendothelial migration without affecting adhesion of the neutrophils to the endothelium. The LLD site-specific mAb VIM12 obviates beta-glucan protection while activation of the LLD by VIM12 cross-linking mimics the beta-glucan response by both preserving endothelial barrier function and reducing neutrophil transendothelial migration. beta-glucan has no direct effect on endothelial cell function in the absence of activated neutrophils. These findings demonstrate that signaling through the CR3 LLD prevents neutrophil-induced loss of endothelial barrier function and reduces diapedesis. This suggests that the LLD may be a suitable target for oligosaccharide-based anti-inflammatory therapeutics.

Chemotaxis, Leukocyte↗
PubMed2004

Antibodies immobilized as arrays to profile protein post-translational modifications in mammalian cells.

Previously, we demonstrated that antibodies printed on a solid support were able to detect protein-protein interaction in mammalian cells. Here we further developed the antibody array system for detecting proteins with various post-translational modifications in mammalian cells. In this novel approach, immunoprecipitated proteins were labeled with fluorescent dye followed by incubation over antibody arrays. Targeted proteins, captured by the antibodies immobilized on PVDF membrane or glass slide, were detected by means of near infrared fluorescent scanner or fluorescent microscopy. To demonstrate the application of the antibody arrays in protein post-translational modifications, we profiled protein tyrosine phosphorylation, ubiquitination, and acetylation in mammalian cells under different conditions. Our results indicate that antibody array technology can provide a powerful means of profiling a large number of proteins with different post-translational modifications in cells.

3T3 Cells↗
PubMed2004

Beta-glucan affects leukocyte navigation in a complex chemotactic gradient.

BACKGROUND: Polymorphonuclear leukocytes (PMNs) must traverse endogenous chemotactic gradients (interleukin 8 [IL-8]) before reaching target chemoattractants (fMLP [N-formylmethionine-leucine-phenylalanine], C5a) produced at a site of bacterial infection. Complement receptor 3 (CR3; CD11b/CD18) contains 2 distinct binding sites, one that mediates adhesion and a lectin-like domain (LLD) that binds polysaccharides of microbial origin. This laboratory previously reported an increase in the chemotactic capacity of PMNs toward fMLP upon ligation of the CR3 LLD with beta-glucan, a CR3 agonist. Current studies sought to determine the effect of beta-glucan on PMN navigation toward other chemoattractants alone and in a competing chemotactic environment. METHODS: Migration was assessed by serum agarose overlay with the use of chambered slides containing or not, beta-glucan. Migration of human PMNs at 37 degrees C for 2 hours was evaluated toward C5a or IL-8 alone and in competing gradients. Selected groups were treated with anti-CR3-blocking antibodies. The number of chemotactic cells was quantified by microscopy. RESULTS: beta-glucan significantly enhanced chemotaxis toward C5a and suppressed that toward IL-8 in a CR3-dependent fashion. In the competing chemotactic gradient assays (C5a vs IL-8), beta-glucan further enhanced migration toward C5a while not affecting that toward IL-8. CONCLUSIONS: beta-glucan selectively upregulates PMN chemotaxis toward C5a while suppressing chemotaxis toward IL-8.

Cell Movement↗
PubMedDate not supplied

Oxygen and the regulation of gene expression in wounds.

Work from Tom Hunt's laboratory first identified wound hypoxia as a potential regulator of the biology of cells participating in tissue repair. Current understanding of the role of oxygen in the regulation of gene expression begins to provide a mechanistic basis for the prediction that oxygen could be a fundamental regulator of wound healing made by the Hunt laboratory. The present article describes the experience of the authors' laboratory in defining the expression of two oxygen-regulated genes, those for the inducible form of nitric oxide synthase and for arginase I in experimental wounds. Observations made regarding these two genes are discussed in the context of the overall regulatory role of oxygen as a phenotypic modulator of inflammatory cells.

Animals↗