TNF Production In Mice Research Articles

Antagonism of the histamine H4 receptor reduces LPS-induced TNF production in vivo

ObjectiveAntagonism of the histamine H4 receptor (H4R) has been shown to be anti-inflammatory in a number of preclinical disease models, however the exact mechanisms behind this are still being uncovered. In vitro, the receptor interacts with TLR and impacts inflammatory mediator production from a number of different cell types. Here it is shown that this interaction also occurs in vivo.Materials and methodsWild-type and H4R deficient BALB/c mice received an i.p. injection of LPS in PBS in conjunction with p.o. JNJ 7777120 or JNJ 28307474 (H4R antagonists). Two hours later blood was collected and TNF was measured.ResultsTwo different H4R antagonists inhibited LPS-induced TNF production in mice and this production was also reduced in H4R-deficient mice. The TNF mRNA analysis showed that the major source of the cytokine was the liver and not blood, and that the H4R antagonist only reduced the expression levels in the liver. Depletion or inactivation of macrophages reduced the TNF levels and eliminated the H4R sensitivity. Treatment with an H4R antagonist also reduced LPS-induced liver injury and blocked LPS-enhanced lung inflammation in mice.ConclusionThe data support an interaction between H4R and TLR activation in vivo that can drive inflammatory responses.Electronic supplementary materialThe online version of this article (doi:10.1007/s00011-013-0612-5) contains supplementary material, which is available to authorized users.

Intestinal epithelial cells as producers but not targets of chronic TNF suffice to cause murine Crohn-like pathology

TNF plays a crucial role in the pathogenesis of Crohn disease. Dysregulated TNF production in mice that bear the genetic deletion of the TNF AU-rich regulatory elements (ARE) (Tnf(ΔARE/+) mice) results in TNF receptor I (TNFRI)-dependent spontaneous Crohn-like pathology. Current concepts consider intestinal epithelial cell (IEC) responses to TNF to be critical for intestinal pathology, but the potential contribution of IEC-derived TNF in disease pathogenesis has not been addressed. In this study we examined whether IEC are sufficient as cellular targets or sources of TNF in the development of intestinal pathology. Using IEC-specific reactivation of a hypomorphic Tnf(ΔAREneo) allele in mice, we show that selective chronic overproduction of TNF by IEC suffices to cause full development of Crohn-like pathology. Epithelial TNF overexpression leads to early activation of the underlying intestinal myofibroblast, a cell type previously identified as a sufficient target of TNF for disease development in the Tnf(ΔARE) model. By contrast, restricted TNFRI expression on IEC although sufficient to confer IEC apoptosis after acute exogenous TNF administration, fails to induce pathology following chronic specific targeting of IEC by endogenous TNF in Tnf(ΔARE/+) mice. Our results argue against IEC being early and sufficient responders to chronic TNF-mediated pathogenic signals and suggest that proinflammatory aberrations leading to chronic TNF production by IEC may initiate pathology in Crohn disease.

Inhibition of colony-stimulating-factor-1 signaling in vivo with the orally bioavailable cFMS kinase inhibitor GW2580.

Colony-stimulating-factor-1 (CSF-1) signaling through cFMS receptor kinase is increased in several diseases. To help investigate the role of cFMS kinase in disease, we identified GW2580, an orally bioavailable inhibitor of cFMS kinase. GW2580 completely inhibited human cFMS kinase in vitro at 0.06 microM and was inactive against 26 other kinases. GW2580 at 1 microM completely inhibited CSF-1-induced growth of mouse M-NFS-60 myeloid cells and human monocytes and completely inhibited bone degradation in cultures of human osteoclasts, rat calvaria, and rat fetal long bone. In contrast, GW2580 did not affect the growth of mouse NS0 lymphoblastoid cells, human endothelial cells, human fibroblasts, or five human tumor cell lines. GW2580 also did not affect lipopolysaccharide (LPS)-induced TNF, IL-6, and prostaglandin E2 production in freshly isolated human monocytes and mouse macrophages. After oral administration, GW2580 blocked the ability of exogenous CSF-1 to increase LPS-induced IL-6 production in mice, inhibited the growth of CSF-1-dependent M-NFS-60 tumor cells in the peritoneal cavity, and diminished the accumulation of macrophages in the peritoneal cavity after thioglycolate injection. Unexpectedly, GW2580 inhibited LPS-induced TNF production in mice, in contrast to effects on monocytes and macrophages in vitro. In conclusion, GW2580's selective inhibition of monocyte growth and bone degradation is consistent with cFMS kinase inhibition. The ability of GW2580 to chronically inhibit CSF-1 signaling through cFMS kinase in normal and tumor cells in vivo makes GW2580 a useful tool in assessing the role of cFMS kinase in normal and disease processes.

1-substituted 4-aryl-5-pyridinylimidazoles: a new class of cytokine suppressive drugs with low 5-lipoxygenase and cyclooxygenase inhibitory potency.

A series of 1-alkyl- or -aryl-4-aryl-5-pyridinylimidazoles (A) were prepared and tested for their ability to bind to a recently discovered protein kinase termed CSBP and to inhibit lipopolysaccharide (LPS)-stimulated TNF production in mice. The kinase, CSBP, appears to be involved in a signaling cascade initiated by a number of inflammatory stimuli and leading to the biosynthesis of the inflammatory cytokines IL-1 and TNF. Two related imidazole classes (B and C) had previously been reported to bind to CSBP and to inhibit LPS-stimulated human monocyte IL-1 and TNF production. The members of the earlier series exhibited varying degrees of potency as inhibitors of the enzymes of arachidonic acid metabolism, PGHS-1 and 5-LO. Several of the more potent CSBP ligands and TNF biosynthesis inhibitors among the present series of N-1-alkylated imidazoles (A) were tested as inhibitors of PGHS-1 and 5-LO and were found to be weak to inactive as inhibitors of these enzymes. One of the compounds, 9 (SB 210313) which lacked measureable activity as an inhibitor of the enzymes of arachidonate metabolism, and had good potency in the binding and in vivo TNF inhibition assays, was tested for antiarthritic activity in the AA rat model of arthritis. Compound 9 significantly reduced edema and increased bone mineral density in this model.

Role of the hypothalamus-pituitary-adrenal axis in the regulation of TNF production in mice. Effect of stress and inhibition of endogenous glucocorticoids.

Glucocorticoids (GC) are known inhibitors of TNF production and are increased by endotoxin (LPS) through a stimulation of the hypothalamus-pituitary-adrenal axis (HPAA), suggesting a feedback mechanism. We tried different approaches to study the role of the HPAA and endogenous GC in the regulation of TNF production. Cyanoketone, a GC synthesis inhibitor, inhibited corticosterone (CS) induction by LPS and increased LPS-induced serum TNF levels. Similar results were obtained by pretreating mice with anticorticotropin-releasing hormone Abs. Administration of adrenocorticotropic hormone increased blood CS and inhibited LPS-induced serum TNF. TNF production by mouse blood stimulated in vitro with LPS was inhibited by addition of CS. Blood from stressed or adrenocorticotropic-releasing hormone-treated mice (in which CS levels are elevated) stimulated ex vivo with LPS produced significantly less TNF than blood from control mice. Normal TNF production was restored by the addition of the GC receptor antagonist RU 38486, indicating a role for the elevated endogenous CS levels in the inhibition of TNF production. These data indicate that the HPAA is a major regulator of TNF production.

Production of tumor necrosis factor induced by synthetic low-toxicity lipid a analog, DT-5461a, is mediated by LPS receptor sites and tyrosine kinase—MAP kinase signaling pathway in murine macrophages

The synthetic low-toxicity lipid A analog DT-5461a induces endogenous TNF production in mice. The activity of TNF so induced is probably the main contributor to the antitumor effect of this compound. In the present study, we investigated the mechanism by which DT-5461a induces TNF production in murine macrophage RAW 264 cells. DT-5461a mimicked the ability of LPS to induce TNF production in a dose-dependent manner. DT-5461a at higher concentrations inhibited specific binding of [ 3H]LPS to the cells and reduced LPS-induced TNF production to the level induced by DT-5461a alone. In addition, DT-5461a, as well as LPS, induced tyrosine phosphorylation of MAP kinases, the early signal transduction pathway of this production. Herbimycin A, an inhibitor of tyrosine kinase, inhibited the LPS- and DT-5461a-induced tyrosine phosphorylation, expression of TNF mRNA, and subsequent TNF secretion. These results suggest that DT-5461a and LPS induce TNF production in murine macrophages through the common receptor sites and the similar early signaling pathway.

Role of the hypothalamus-pituitary-adrenal axis (HPAA) in the regulation of TNF production in mice

Glucocorticoids (CC) are known inhibitors of TNF production and are increased by endotoxin (LPS) through a stimulation of the hypothalamus-pituitary-adrenal axis (HPAA), suggesting a feedback mechanism. We tried different approaches to study the role of the HPAA and endogenous CC in the regulation of TNF production. Cyanoketone, a CC synthesis inhibitor, inhibited corticosterone (CS) induction by LPS and increased LPS-induced serum TNF levels. Similar results were obtained by pretreating mice with anticorticotropin-releasing hormone Abs. Administration of adrenocorticotropic hormone increased blood CS and inhibited LPS-induced serum TNF. TNF production by mouse blood stimulated in vitro with LPS was inhibited by addition of CS. Blood from stressed or adrenocorticotropic-releasing hormone-treated mice (in which CS levels are elevated) stimulated ex vivo with LPS produced significantly less TNF than blood from control mice. Normal TNF production was restored by the addition of the CC receptor antagonist RU 38486, indicating a role for the elevated endogenous CS levels in the inhibition of TNF production. These data indicate that the HPAA is a major regulator of TNF production. The journal of Immunology, 1995,155: 3552-3555.

P-564 - Effects of aprotinin on TNF production in mice with endotoxin-induced shock.

P-564 - Effects of aprotinin on TNF production in mice with endotoxin-induced shock.

Patent Evaluation: Lipid-A analogues as inhibitors of lipopolysaccharide-induced TNF production

SummarySummaryNovelty: Analogues of lipid A useful as anti-endotoxic drugs for the treatment of septic shock and lipopolysaccharide (LPS)-mediated activation of viral infection (HIV, herpes or influenza) are disclosed.Biology: The compounds were assayed for their ability to inhibit LPS-induced production of tumour necrosis factor and IL-1β both in vitro and in vivo. One compound effectively inhibited TNF production in mice with an ED50 value of 5.4μg/mouse.Chemistry: The synthesis of the compounds are described in three examples including that of analogue B318.

In vivo production of exotoxin A and its role in endogenous Pseudomonas aeruginosa septicemia in mice

We have examined the production of Pseudomonas aeruginosa exotoxin A (ETA) and its role in endogenous bacteremia in mice. Mice given P. aeruginosa D4 orally died of bacteremia between days 10 and 13 following cyclophosphamide-induced leukocytopenia. In this model, serum endotoxin was detected beginning on day 7 by the Limulus assay and P. aeruginosa was cultured from blood beginning on day 9. ETA and tumor necrosis factor alpha (TNF) were also detected in serum by enzyme-linked immunosorbent assay beginning on day 9. Purified ETA did not stimulate the production of TNF in normal mice primed with a synthetic derivative of muramyl dipeptide in the absence of endotoxin. However, ETA enhanced and primed endotoxin-induced TNF production in mice. The mortality rate of mice given ETA mutant PAO-PRI (5.0%) was significantly lower than that of mice given the parent strain (78.8%). These data indicate that ETA may be an important factor in the occurrence of P. aeruginosa bacteremia and/or the death of mice. Also, ETA may be responsible for enhancing the production of a lethal dose of TNF in the presence of endotoxin in P. aeruginosa bacteremia.