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RAW264.7 Macrophage Cell Line (raw264.7 + macrophage_cell_line)
Selected AbstractsInterferon-, and lipopolysaccharide regulate the expression of Nramp2 and increase the uptake of iron from low relative molecular mass complexes by macrophagesFEBS JOURNAL, Issue 22 2000S. L. Wardrop The natural resistance associated macrophage protein 2 (Nramp2) is a transporter that is involved in iron (Fe) uptake from transferrin (Tf) and low molecular mass Fe complexes. Here we describe the effect of the inflammatory mediators interferon-, (IFN-,) and lipopolysaccharide (LPS) on the expression of Nramp2 mRNA and Fe uptake by cells of the macrophage lineage. After incubation of the RAW264.7 macrophage cell line with LPS there was a sevenfold increase in the expression of the 2.3 kb Nramp2 mRNA transcript when compared with the control, but little effect on the Nramp2 3.1 kb transcript. These results indicate differential regulation of the two transcripts. Treatment with LPS resulted in an increase in 59Fe uptake from 59Fe,nitrilotriacetic acid, while transferrin receptor (TfR) mRNA levels and 59Fe uptake from 59Fe,Tf were decreased. Paradoxically, at the same time, an increase in iron regulatory protein (IRP)1 RNA-binding activity was observed. Incubation with IFN-, (50 U·mL,1) resulted in a marked decrease in TfR mRNA levels but had no effect on Nramp2 mRNA expression. Exposure of RAW264.7 cells to both IFN-, and LPS resulted in a fourfold increase in the Nramp2 2.3-kb transcript and a four to fivefold decrease in the 3.1-kb transcript when compared with the control. Furthermore, there was a decrease in TfR mRNA levels despite an increase in IRP1 RNA-binding activity and a marked increase in inducible nitric oxide synthase mRNA expression. Hence, TfR and Nramp2 mRNA expression did not appear to be regulated in a concerted manner. Similar responses to those found above for RAW264.7 cells were also observed in the J774 macrophage cell line and also for primary cultures of mouse peritoneal macrophages. These results are of interest as the TfR and Nramp2 are thought to act together during Fe uptake from Tf. This is the first report to demonstrate regulation of the Nramp2 mRNA transcripts by inflammatory mediators. [source] A Macrophage Cell Model for Selective Metalloproteinase Inhibitor DesignCHEMBIOCHEM, Issue 13 2008Faith E. Jacobsen Abstract The desire to inhibit zinc-dependent matrix metalloproteinases (MMPs) has, over the course of the last 30 years, led to the development of a plethora of MMP inhibitors that bind directly to the active-site metal. With one exception, all of these drugs have failed in clinical trials, due to many factors, including an apparent lack of specificity for MMPs. To address the question of whether these inhibitors are selective for MMPs in a biological setting, a cell-based screening method is presented to compare the relative activities of zinc, heme iron, and non-heme iron enzymes in the presence of these compounds using the RAW264.7 macrophage cell line. We screened nine different zinc-binding groups (ZBGs), four established MMP inhibitors (MMPis), and two novel MMP inhibitors developed in our laboratory to determine their selectivities against five different metalloenzymes. Using this model, we identified two nitrogen donor compounds,2,2,-dipyridylamine (DPA) and triazacyclononane (TACN),as the most selective ZBGs for zinc metalloenzyme inhibitor development. We also demonstrated that the model could predict known nonspecific interactions of some of the most commonly used MMPis, and could also give cross-reactivity information for newly developed MMPis. This work demonstrates the utility of cell-based assays in both the design and the screening of novel metalloenzyme inhibitors. [source] A nitric oxide (NO)-releasing derivative of gabapentin, NCX 8001, alleviates neuropathic pain-like behavior after spinal cord and peripheral nerve injuryBRITISH JOURNAL OF PHARMACOLOGY, Issue 1 2004Wei-Ping Wu Nitric oxide (NO) participates, at least in part, to the establishment and maintenance of pain after nerve injury. Therefore, drugs that target the NO/cGMP signaling pathway are of interest for the treatment of human neuropathic pain. Various compounds endowed with NO-releasing properties modulate the expression and function of inducible nitric oxide synthase (iNOS), the key enzyme responsible for sustained NO production under pathological conditions including neuropathic pain. With this background, we synthesized a new chemical entity, [1-(aminomethyl)cyclohexane acetic acid 3-(nitroxymethyl)phenyl ester] NCX8001, which has a NO-releasing moiety bound to gabapentin, a drug currently used for the clinical management of neuropathic pain. We examined the pharmacological profile of this drug with respect to its NO-releasing properties in vitro as well as to its efficacy in treating neuropathic pain conditions (allodynia) consequent to experimental sciatic nerve or spinal cord injuries. NCX8001 (1,30 ,M) released physiologically relevant concentrations of NO as it induced a concentration-dependent activation of soluble guanylyl cyclase (EC50=5.6 ,M) and produced consistent vasorelaxant effects in noradrenaline-precontracted rabbit aortic rings (IC50=1.4 ,M). NCX8001, but not gabapentin, counteracted in a concentration-dependent fashion lipopolysaccharide-induced overexpression and function of iNOS in RAW264.7 macrophages cell line. Furthermore, NCX8001 also inhibited the release of tumor necrosis factor alpha (TNF,) from stimulated RAW264.7 cells. NCX8001 (28,280 ,mol kg,1, i.p.) reduced the allodynic responses of spinal cord injured rats in a dose-dependent fashion while lacking sedative or motor effects. In contrast, gabapentin (170,580 ,mol kg,1, i.p.) resulted less effective and elicited marked side effects. NCX8001 alleviated the allodynia-like responses of rats to innocuous mechanical or cold stimulation following lesion of the sciatic nerve. This effect was not shared by equimolar doses of gabapentin. Potentially due to the slow releasing kinetics of NO, NCX8001 alleviated pain-like behaviors in two rat models of neuropathic pain in a fashion that is superior to its parent counterpart gabapentin. This new gabapentin derivative, whose mechanism deserves to be explored further, offers new hopes to the treatment of human neuropathic pain. British Journal of Pharmacology (2004) 141, 65,74. doi:10.1038/sj.bjp.0705596 [source] Effects of phenylethanoid glycosides from Digitalis purpurea L. on the expression of inducible nitric oxide synthaseJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 7 2005Jae Wook Oh We have isolated four different phenylethanoid glycosides (purpureaside A, desrhamnosyl acteo-side, calceolarioside B and plantainoside D) from the leaves of Digitalis purpurea (foxglove). The effects of these glycosides on activator protein-1 (AP-1)-mediated inducible nitric oxide synthase (iNOS) gene expression in the Raw264.7 macrophage cell line have been studied. Of these four glycosides, purpureaside A potently inhibited iNOS induction by lipopolysaccharide (LPS). Increase in iNOS mRNA by LPS was completely suppressed by purpureaside A. Purpureaside A did not significantly affect LPS-inducible nuclear factor- kB (NF- kB) activation or the nuclear translocation of p65. Moreover, a reporter gene assay using AP-1 specific luciferase reporter revealed that the enhanced activity of AP-1 by LPS was completely abolished in cells treated with purpureaside A. These results demonstrated that purpureaside A inhibited LPS-inducible iNOS expression in macrophages through the suppression of AP-1, but not of NF- kB. [source] | |||||||||||||