Transgenic mice overexpressing human ALOX15 under the control of the aP2 promoter are partly protected in the complete Freund's adjuvant-induced paw inflammation model.

BACKGROUND, OBJECTIVES AND DESIGN: Arachidonic acid 15-lipoxygenase (ALOX15) has been implicated in the pathogenesis of inflammatory diseases but since pro- and anti-inflammatory roles have been suggested, the precise function of this enzyme is still a matter of discussion. To contribute to this discussion, we created transgenic mice, which express human ALOX15 under the control of the activating protein 2 promoter (aP2-ALOX15 mice) and compared the sensitivity of these gain-of-function animals in two independent mouse inflammation models with Alox15-deficient mice (loss-of-function animals) and wildtype control animals. MATERIALS AND METHODS: Transgenic aP2-ALOX15 mice were tested in comparison with Alox15 knockout mice (Alox15-/-) and corresponding wildtype control animals (C57BL/6J) in the complete Freund's adjuvant induced hind-paw edema model and in the dextran sulfate sodium induced colitis (DSS-colitis) model. In the paw edema model, the degree of paw swelling and the sensitivity of the inflamed hind-paw for mechanic (von Frey test) and thermal (Hargreaves test) stimulation were quantified as clinical readout parameters. In the dextran sodium sulfate induced colitis model the loss of body weight, the colon lengths and the disease activity index were determined. RESULTS: In the hind-paw edema model, systemic inactivation of the endogenous Alox15 gene intensified the inflammatory symptoms, whereas overexpression of human ALOX15 reduced the degree of hind-paw inflammation. These data suggest anti-inflammatory roles for endogenous and transgenic ALOX15 in this particular inflammation model. As mechanistic reason for the protective effect downregulation of the pro-inflammatory ALOX5 pathways was suggested. However, in the dextran sodium sulfate colitis model, in which systemic inactivation of the Alox15 gene protected female mice from DSS-induced colitis, transgenic overexpression of human ALOX15 did hardly impact the intensity of the inflammatory symptoms. CONCLUSION: The biological role of ALOX15 in the pathogenesis of inflammation is variable and depends on the kind of the animal inflammation model.

Inhibition of oxygen-induced ischemic retinal neovascularization with adenoviral 15-lipoxygenase-1 gene transfer via up-regulation of PPAR-γ and down-regulation of VEGFR-2 expression.

15-lipoxygenase-1 (15-LOX-1) plays an important role in angiogenesis, but how it works still remains a controversial subject. The aims of our study are focused on determining whether or not 15-LOX-1 inhibiting oxygen-induced ischemic retinal neovascularization (RNV) and the underlying regulatory mechanism involving of 15-LOX-1, peroxisome proliferator-activated receptor γ (PPAR-γ) and vascular endothelial growth factor receptor 2 (VEGFR-2) in oxygen-induced retinopathy (OIR). Recombinant adenoviral vectors that expressing the 15-LOX-1 gene (Ad-15-LOX-1-GFP) or the green fluorescence protein gene (Ad-GFP) were intravitreous injected into the OIR mice at postnatal day 12 (P12), the mice were sacrificed 5 days later (P17). Retinal 15-LOX-1 expression was significantly increased at both mRNA and protein levels after 15-LOX-1 gene transfer. Immunofluorescence staining of retinal sections revealed 15-LOX-1 expression was primarily in the outer plexiform layer (OPL), inner nuclear layer (INL) and ganglion cell layer (GCL) retina. Meanwhile, RNV was significantly inhibited indicated by fluorescein retinal angiography and quantification of the pre-retinal neovascular cells. The expression levels of PPAR-γ were significantly up-regulated while VEGFR-2 were significantly down-regulated both in mRNA and protein levels. Our results suggested 15-LOX-1 gene transfer inhibited RNV in OIR mouse model via up-regulation of PPAR-γ and further down-regulation of VEGFR-2 expression. This could be a potentially important regulatory mechanism involving 15-LOX-1, PPAR-γ and VEGFR-2 during RNV in OIR. In conclusion, 15-LOX-1 may be a new therapeutic target for treating neovascularization diseases.

Synergistic effect of 15-lipoxygenase 2 and radiation in killing head-and-neck cancer.

We previously demonstrated that 15-LOX-2 is significantly reduced in head and neck carcinoma and restoration of 15-LOX-2 expression results in tumor inhibition in HNC. The aim of this study is to evaluate 15-LOX-2 as a candidate for targeted radiotherapy. Molecular subcloning was performed to create a radiation-inducible 15-LOX-2 expression vector in which the full-length 15-LOX-2 cDNA was inserted downstream the recombinant Egr-1 promoter. The radiation-induced downregulations of 15-LOX-2 protein (twofold up) and its main metabolite 15S-HETE (threefold up) were observed in HNC cells transfected with the 15-LOX-2 expression vector after 4 Gy of radiation. Radiation-induced upregulation of 15-LOX-2 resulted in significant induction of apoptosis in HNC cells. Furthermore, survival colony formation was significantly reduced by 4 Gy in the HNC cells containing the 15-LOX-2 expression vector compared with the controls. Radiation-induced upregulation of 15-LOX-2 results in significant induction of apoptosis and enhances killing effect of radiotherapy in HNC. In addition, exogenous addition of 15S-HETE at high concentrations (>/=10 muM) but not at low concentrations induced upregulation of its endogenous ligand PPARgamma. In conclusion, synergistic effect between radiation and 15-LOX-2 was observed in killing HNC. 15-LOX-2 may be a potential target in radiation-targeted therapy of HNC. The 15-LOX-2 inhibition may not be PPARgamma dependent.