Angel Montero

Human 15-Lipoxygenase gene promoter: analysis and identification of DNA binding sites for IL-13-induced regulatory factors in monocytes

In order to study the transcriptional control of 15-LO expression, we have cloned and sequenced the human 15-LO promoter region. The 15-LO promoter is associated with a CpG island at the 5′-end of the gene, and sequence analysis reveals putative Sp1 and Ap2 binding site/s and absence of TATA or CAAT motifs. Transcription is initiated at one major site. Using deletion constructs, we have defined an active promoter region of 1056 bp. Gel-shift assays revealed that transcriptional factor(s) induced only in response to IL-13 treatment of human peripheral blood monocytes bind to the 15-LO promoter DNA. Two regions, DP1 (−140 to −92 bp) and DP2 (−353 to −304 bp) of the promoter were essential for transcription in HeLa cells and human peripheral monocytes. Hela nuclear extracts contained a specific nuclear factor(s) binding to 15-LO promoter DNA which are distinct from those derived from IL-13-treated human peripheral monocyte nuclear extracts. In addition, fluorescent in situ hybridization (FISH) results refined the previous localization of 15-LO to human chromosome 17p13.3.

Reciprocal Regulation of LTA4 Hydrolase Expression in Human Monocytes by γ-Interferon and Interleukins 4 and 13: Potential Relevance to Leukotriene Regulation in Glomerular Disease

Background/Aims: Leukotriene A₄ (LTA₄) hydrolase catalyzes the final step in the synthesis of leukotriene B₄ (LTB₄). TH-1- and TH-2-derived cytokines may regulate LTB₄ synthesis by monocytes through their actions on the expression of LTA₄ hydrolase. Methods: Freshly isolated monocytes were incubated with pro- and anti-inflammatory cytokines for 36 h. mRNA expression was determined by Northern blot, protein expression was determined by Western blot and LTB₄ synthesis was determined by ELISA. Results: Interferon-γ (a TH-2-derived cytokine) increased significantly LTA₄ hydrolase mRNA expression, whereas interleukin (IL)-4 and IL-13 (both TH-2-derived cytokines) decreased LTA₄ hydrolase mRNA expression in these cells. The same effects were seen on the expression of immunoreactive LTA₄ hydrolase after incubating the monocytes with either TH-1- or TH-2-derived cytokines. The monocyte-derived cytokine IL-1β did not show any significant effect on LTA₄ hydrolase mRNA expression. When LTB₄ release was measured, both IL-1β and interferon-γ significantly increased LTB₄ production by monocytes, while TH-2 cytokines (IL-4 and IL-13) decreased it. Conclusion: The opposing effects of TH-1- and TH-2-derived cytokines on the expression of LTA₄ hydrolase mRNA may regulate LTB₄ synthesis by monocytes during inflammation.

15-lipoxygenase in glomerular inflammation

Glomerulonephritis is a significant factor fueling the rapid increase in the population of patients with end-stage renal disease. Novel therapeutic strategies targeting specific mechanisms of glomerular destruction are the most reasonable approaches to arrest ongoing injury. In this review, we summarize some of our results obtained in our effort to characterize the role of 15-lipoxygenase activation as one of the mechansisms operative during the early, prefibrotic stage of glomerular immune injury. We also summarize the effects of cytokines released during these processes, as well as the activation by aspirin of the synthesis of 15-R-HETE (see text). Finally, we will propose a clinical approach to this group of disorders, based on emerging concepts of the pathophysiology of glomerulonephritis from our work and that of several other investigators.

LTA4 Hydrolase Expression During Glomerular Inflammation: Correlation of Immunohistochemical Localization with Cytokine Regulation

Leukotriene B4 (LTB4) is the most potent chemotactic substance described for polymorphonuclear leukocytes (PMN) and it also increases PMN aggregation and adhesion to endothelial cells (Samuelson et al., 1987). Glomerular LTB4 synthesis is enhanced early in the course of several forms of glomerular injury, particularly those characterized by PMN or macrophage infiltration (for review, see Badr, 1992). Most of the studies performed agree on the transient pattern of LTB4 production following glomerular injury (Badr, 1992, Katoh et al., 1994). LTB4 production is enhanced during the first 24 hours of injury, and after this period its levels are undetectable in most proliferative and non-proliferative glomerulopathies.

Identification and characterization of an enhancer sequence in the promoter region of human 15-lipoxygenase (15-LO) gene

Lipoxygenases are lipid-peroxidating enzymes that are implicated in the pathogenesis of a variety of inflammatory disorders 1-3, membrane remodeling 4-6 and atheroma formation 7-9. Formation of 15-(S)-hydroxyeicosatetraenoic acid [15-(S)-HETE] and lipoxin (LX) A4 in human leukocytes, mediated by 15-LO dependent catalysis of arachidonic acid, likely represents a component of endogenous pro- and anti-inflammatory influences that ultimately regulate the extent and severity of inflammatory reactions 1-4, 10.

Inhibition of 5-lo Activating Protein (Flap) Activity Decreases Proteinuria in Streptozotocin(Stz)-induced Diabetic Rats.

Proteinuria is a major cause of morbidity in patients with glomerular disease. Nonselective protein filtration is now recognized as a significant contributing factor to the tubulo-interstitial injury which underlies and often predicts progressive renal insufficiency in patients with primary glomerular diseases. Increasing recognition of the mechanisms whereby proteinuria leads to tubule injury confirms the well-known clinical observation that the magnitude of proteinuria at diagnosis is a reliable predictor of prognosis in patients with glomerulonephritis and diabetes. For this reason, the search for safe and effective anti-proteinuric agents which do not reduce the glomerular filtration rate represents an urgent and critical need for physicians involved in the care of patients with glomerular disorders.