Deuterated polyunsaturated fatty acids inhibit photoirradiation-induced lipid peroxidation in lipid bilayers.

Lipid peroxidation (LPO) plays a key role in many age-related neurodegenerative conditions and other disorders. Light irradiation can initiate LPO through various mechanisms and is of importance in retinal and dermatological pathologies. The introduction of deuterated polyunsaturated fatty acids (D-PUFA) into membrane lipids is a promising approach for protection against LPO. Here, we report the protective effects of D-PUFA against the photodynamically induced LPO, using illumination in the presence of the photosensitizer trisulfonated aluminum phthalocyanine (AlPcS3) in liposomes and giant unilamellar vesicles (GUV), as assessed in four experimental models: 1) sulforhodamine B leakage from liposomes, detected with fluorescence correlation spectroscopy (FCS); 2) formation of diene conjugates in liposomal membranes, measured by absorbance at 234 nm; 3) membrane leakage in GUV assessed by optical phase-contrast intensity observations; 4) UPLC-MS/MS method to detect oxidized linoleic acid (Lin)-derived metabolites. Specifically, in liposomes or GUV containing H-PUFA (dilinoleyl-sn-glycero-3-phosphatidylcholine), light irradiation led to an extensive oxidative damage to bilayers. By contrast, no damage was observed in lipid bilayers containing 20% or more D-PUFA (D2-Lin or D10-docosahexanenoic acid). Remarkably, addition of tocopherol increased the dye leakage from liposomes in H-PUFA bilayers compared to photoirradiation alone, signifying tocopherol's pro-oxidant properties. However, in the presence of D-PUFA the opposite effect was observed, whereby adding tocopherol increased the resistance to LPO. These findings suggest a method to augment the protective effects of D-PUFA, which are currently undergoing clinical trials in several neurological and retinal diseases that involve LPO.

[Inflammatory metabolites of arahidonic acid in tear fluid in UV-induced corneal damage]. / Vospalitel'nye metabolity arakhidonovoi kisloty v sleznoi zhidkosti pri indutsirovannom ul'trafioletovym izlucheniem povrezhdenii rogovitsy.

The ultraviolet (UV) B-induced damage of the eye surface of experimental animals (rabbits) includes loss of corneal epithelium, apoptosis of keratocytes and stromal edema. These changes are accompanied by clinically and histologically manifested corneal inflammation, neutrophil infiltration, and exudation of the anterior chamber of the eye. According to mass spectrometric analysis, UV-induced corneal damage is associated with pronounced changes in the lipid composition of tears, including a decrease in the amount of arachidonic acid and prostaglandin E2 and an increase in the concentrations of prostaglandin D2 and its derivative 15d-PGJ2. In addition, it is accompanied by an alteration in the levels of hydroxyeicosate tetraenic acid derivatives, namely upregulation of 12-HETE and downregulation of 5-HETE. The revealed changes indicate the activation of metabolic pathways involving 5-lipoxygenase, 12-lipoxygenase, cyclooxygenase 1 and 2, and prostaglandin-D-synthase. These findings contribute to understanding mechanisms of UV-induced keratitis and point on feasibility of selective anti-inflammatory therapy for improving corneal regeneration after iatrogenic UV damage.

Antiinflammatory Effect of Rosiglitazone via Modulation of mRNA Stability of Interleukin 10 and Cyclooxygenase 2 in Astrocytes.

Investigation of molecular mechanisms of proinflammatory stimuli signaling in astrocytes is important for understanding their role in pathogenesis of central nervous system diseases as well as in functioning of the innate immunity system in non-immune cells. Here we show that lipopolysaccharide (LPS) stimulation of primary rat astrocytes led to conventional inflammatory response: increase in both proinflammatory (tumor necrosis factor, TNFα; prostaglandin E2, PGE2) and antiinflammatory marker (interleukin 10, IL-10) levels. The protein level of cyclooxygenase 2 (COX-2) was also increased. Rosiglitazone strengthened LPS-induced mRNA expression of COX-2 and IL-10 but not TNFα. Rosiglitazone is an agonist of nuclear receptor PPARγ, but its impact on IL-10 expression was not influenced by a PPARγ antagonist, GW9662, suggesting PPARγ-independent effect of rosiglitazone. The degradation of mRNA is one of the steps of inflammation regulation and might be affected by small molecules. In experiments with actinomycin D, we found that mRNA half-lives of IL-10, COX-2, and TNFα in naive astrocytes were 70, 44, and 19 min, respectively. LPS stimulation caused 2-fold increase in IL-10 and COX-2 mRNA decay rates, whereas addition of rosiglitazone restored them to the initial level. TNFα decay rate was not changed by these stimulations. This suggests that mRNA decay rate could be regulated by small molecules. Moreover, rosiglitazone could be used as a substance stimulating the resolution of inflammation without influence on proinflammatory signals. These results open new perspectives in the search for inflammation resolution modulators.

Regulation of cyclooxygenase 2 expression by agonists of PPAR nuclear receptors in the model of endotoxin tolerance in astrocytes.

Endotoxin tolerance (ET) represents a state of an altered immune response induced by multiple stimulations of a cell, a tissue, or an organism with lipopolysaccharide. Characteristics of ET include downregulation of induction of proinflammatory genes (TNFα, IL6, and others) and enhancement of induction of antiinflammatory genes (IL10, TGFß). ET generally has protective functions; nevertheless, it might result in a state of innate immune deficiency and cause negative outcomes. A current issue is the search for the mechanisms controlling the level of inflammation in the course of endotoxin tolerance. In this work, we investigated the change in cyclooxygenase 2 (Cox2) expression in the model of endotoxin tolerance in astrocytes and analyzed the possibility of regulating this process applying nuclear receptor PPAR agonists. Our results indicate that: 1) endotoxin tolerance can be induced in astrocytes and results in TNFα and Cox2 mRNA induction decrease upon secondary stimulation; 2) tolerance is revealed on the level of TNFα release and Cox2 protein expression; 3) PPAR agonists GW7647, L-165041, and rosiglitazone control Cox2 mRNA expression levels under conditions of endotoxin tolerance. In particular, rosiglitazone (a PPARγ agonist) induces Cox2 mRNA expression, while GW7647 (a PPARα agonist) and L-165041 (a PPARß agonist) suppress the expression. Our results demonstrate that Cox2 can be up- and downregulated during endotoxin tolerance in astrocytes, and PPAR agonists might be effective for controlling this target under conditions of multiple proinflammatory stimulations of brain tissues with endotoxin.