Macrophage activation induces formation of the anti-inflammatory lipid cholesteryl-nitrolinoleate.

Nitroalkene derivatives of fatty acids act as adaptive, anti-inflammatory signalling mediators, based on their high-affinity PPARgamma (peroxisome-proliferator-activated receptor gamma) ligand activity and electrophilic reactivity with proteins, including transcription factors. Although free or esterified lipid nitroalkene derivatives have been detected in human plasma and urine, their generation by inflammatory stimuli has not been reported. In the present study, we show increased nitration of cholesteryl-linoleate by activated murine J774.1 macrophages, yielding the mononitrated nitroalkene CLNO2 (cholesteryl-nitrolinoleate). CLNO2 levels were found to increase approximately 20-fold 24 h after macrophage activation with Escherichia coli lipopolysaccharide plus interferon-gamma; this response was concurrent with an increase in the expression of NOS2 (inducible nitric oxide synthase) and was inhibited by the (*)NO (nitric oxide) inhibitor L-NAME (N(G)-nitro-L-arginine methyl ester). Macrophage (J774.1 and bone-marrow-derived cells) inflammatory responses were suppressed when activated in the presence of CLNO2 or LNO2 (nitrolinoleate). This included: (i) inhibition of NOS2 expression and cytokine secretion through PPARgamma and *NO-independent mechanisms; (ii) induction of haem oxygenase-1 expression; and (iii) inhibition of NF-kappaB (nuclear factor kappaB) activation. Overall, these results suggest that lipid nitration occurs as part of the response of macrophages to inflammatory stimuli involving NOS2 induction and that these by-products of nitro-oxidative reactions may act as novel adaptive down-regulators of inflammatory responses.

Nitrated lipids decompose to nitric oxide and lipid radicals and cause vasorelaxation.

Nitric oxide-derived oxidants such as nitrogen dioxide and peroxynitrite have been receiving increasing attention as mediators of nitric oxide toxicity. Indeed, nitrated and nitrosated compounds have been detected in biological fluids and tissues of healthy subjects and in higher yields in patients under inflammatory or infectious conditions as a consequence of nitric oxide overproduction. Among them, nitrated lipids have been detected in vivo. Here, we confirmed and extended previous studies by demonstrating that nitrolinoleate, chlolesteryl nitrolinoleate, and nitrohydroxylinoleate induce vasorelaxation in a concentration-dependent manner while releasing nitric oxide that was characterized by chemiluminescence-and EPR-based methodologies. As we first show here, diffusible nitric oxide production is likely to occur by isomerization of the nitrated lipids to the corresponding nitrite derivatives that decay through homolysis and/or metal ion/ascorbate-assisted reduction. The homolytic mechanism was supported by EPR spin-trapping studies with 3,5-dibromo-4-nitrosobenzenesulfonic acid that trapped a lipid-derived radical during nitrolinoleate decomposition. In addition to provide a mechanism to explain nitric oxide production from nitrated lipids, the results support their role as endogenous sources of nitric oxide that may play a role in endothelium-independent vasorelaxation.

Effects in post-menopausal women of transdermal estrogen associated with progestin upon the removal from the plasma of a microemulsion that resembles low-density lipoprotein (LDL).

OBJECTIVE: To evaluate the effects of transdermal estradiol and medroxyprogesterone acetate (MPA) treatment on the removal from the plasma of a cholesterol-rich microemulsion (LDE) that roughly resembles low-density lipoprotein (LDL) structure and that binds to LDL receptors. METHODS: Ten healthy post-menopausal women were studied before and after 3-month treatment with transdermal estradiol in the following dosages administered every 3.5 days: 25, 50, 50, 100, 100, 50, 50 and 25 microg. From the 15th to the 21st day and from the 22nd to the 28th day of estrogen treatment, respectively, 10 and 5 mg q.d. MPA per oral were associated to the transdermal estrogen. The emulsion labeled with 14C-cholesteryl oleate was injected after 12 h fasting and its fractional catabolic rate (FCR) was calculated from the plasma decaying curves of the isotope. RESULTS: Treatment reduced LDL-cholesterol levels by 8% only (149.0 +/- 36.0 mg/dl, 138.0 +/- 27.0 mg/dl; P = 0.046), but the FCR of LDE expressed in medians (25%; 75%) increased from 0.0054 (0.003; 0.052) h(-1) to 0.021 (0.009; 0.10) h(-1), P = 0.002. CONCLUSION: The association used in this study so as to mimic the increasing-decreasing pattern of the hormonal ovarian production reduced modestly LDL-cholesterol levels but pronouncedly increased the lipoprotein removal as tested by LDE FCR.

Inhibitory effect of flavonoids on low-density lipoprotein peroxidation catalyzed by mammalian 15-lipoxygenase.

Lipoxygenase-dependent low-density lipoprotein (LDL) oxidation is believed to be involved in atherogenesis. Inhibition of lipoxygenase-induced lipid peroxidation might, therefore, be an important mode to suppress the development of atherosclerosis. Because dietary antioxidants inhibit LDL oxidation in vitro and their intake is inversely associated with coronary heart diseases, we compared the inhibitory effect of three typical flavonoids-quercetin, epicatechin, and flavone-with alpha-tocopherol and ascorbic acid against human LDL oxidation catalyzed by mammalian 15-lipoxygenase. The oxidative modification of LDL was monitored by measurement of cholesteryl ester hydroperoxide (CE-OOH) formation and consumption of antioxidants by using HLPC. Quercetin and epicatechin were the strongest inhibitors of LDL oxidation catalyzed by 15-lipoxygenase; ascorbic acid was an effective inhibitor in the first 3 h of oxidation; and fivefold alpha-tocopherol-enriched LDL showed a partial inhibition of CE-OOH formation only after 4-6 h of incubation. Flavone had no effect. Quercetin, ascorbic acid, and alpha-tocopherol were consumed in the first 3 h of incubation. Consumption of LDL alpha-tocopherol was partially inhibited by ascorbic acid and quercetin, whereas epicatechin and flavone were without effect. These results emphasize the inhibitory effect of the flavonoids quercetin and epicatechin on 15-lipoxygenase-mediated LDL lipid peroxidation. At similar concentrations, they are stronger antioxidants than ascorbic acid, alpha-tocopherol, and flavone.

[Immunoregulation of tumor growth in a murine model]. / Inmunorregulación del crecimiento tumoral en un sistema murino.

Different spleen and tumor cell factors modifying tumoral and metastatic growth were studied. Spleen cell culture supernatants (SCS) from small and large tumor-bearing mice enhanced tumor growth. After tumor surgery, tumor enhancement was only mediated by supernatants from large tumor resected mice. Tumor facilitation and angiogenic response were mediated by the same supernatants; different fractions for these two activities were characterized. T and non-T cells, depending on tumor burden, were responsible for the enhancing activity; but angiogenesis depended only on T cells. While augmentation of metastatic spread was produced by tumor antigens (soluble tumor extracts, tumor-cell supernatants, formolized tumor cells), primary tumor development was not modified by tumor-cell supernatants. Increased incidence of metastases was also mediated by SCS from tumor resected mice which had previously been inoculated with tumor antigens. Immune status of tumor-resected mice was evaluated by delayed-type hypersensitivity reaction. Tumor cell membranes enriched with cholesterol-hemisuccinate were able to increase anti-tumor immune response.