Studies on the biological effects of ozone: 10. Release of factors from ozonated human platelets.

In a previous work we have shown that heparin, in the presence of ozone (O3), promotes a dose-dependent platelet aggregation, while after Ca2+ chelation with citrate, platelet aggregation is almost negligible. These results led us to think that aggregation may enhance the release of platelet components. We have here shown that indeed significantly higher amount of platelet-derived growth factor (PDGF), transforming growth factor beta1 (TGF-beta1) and interleukin-8 (IL-8) are released in a dose-dependent manner after ozonation of heparinised platelet-rich plasma samples. These findings may explain the enhanced healing of torpid ulcers in patients with chronic limb ischemia treated with O3 autohaemoteraphy (O3-AHT)

Studies on the biological effects of ozone: 11. Release of factors from human endothelial cells.

BACKGROUND: Empirical observations have shown that ozonated autohemotherapy markedly improves the symptoms of chronic limb ischemia (muscular pain at rest, intermittent claudication, etc) in atherosclerotic patients, but mechanisms of action remain unclear. AIMS: Human endothelial cells (HUVECs) are known to release nitrogen monoxide (NO) and we investigated the biological effects of human ozonated serum on HUVECs in culture. METHODS: We assessed the relevance of peroxidation, the release of NO as nitrite and of three classical cytokines. RESULTS: The treatment of HUVECs with ozonated serum yields a dose dependent increase of thiobarbituric acid reactive substances (TBARS) and of hydrogen peroxide (H2O2) and a decrease of protein thiol groups (PTG). Concomitantly, in comparison to either the control or the oxygenated sample, there is a significant and steady increase of nitric oxide (NO) production; this is markedly enhanced by the addition of L-arginine (20 microM) and inhibited in the presence of the NO inhibitor, L-NAME (20 mM). The main mediator of ozone action is H2O2 as it has been shown either after its direct measurement or by the addition of 20, 40 and 100 microM. Moreover, during 24 hours incubation we have investigated the production of endothelin 1 (ET-1), E-selectin and Interleukin 8 (IL-8) and it appears that ozonation enhances IL-8, inhibits E-selectin and hardly modifies ET-1 production. CONCLUSIONS: It appears that reinfusion of ozonated blood, by enhancing release of NO, may induce vasodilation in ischemic areas and reduce hypoxia

Studies on the biological effects of ozone: 8. Effects on the total antioxidant status and on interleukin-8 production.

Ozone (O3) is a controversial gas because, owing to its potent oxidant properties, it exerts damaging effects on the respiratory tract and yet it has been used for four decades as a therapy. While the disinfectant activity of O3 is understandable, it is less clear how other biological effects can be elicited in human blood with practically no toxicity. On the other hand plasma and cells are endowed with a powerful antioxidant system so that a fairly wide range of O3 concentrations between 40 and 80 microg/ml per gram of blood (approximately 0.83-1.66 mM) are effective but not deleterious. After blood ozonation total antioxidant status (TAS) and plasma protein thiol groups (PTG) decrease by 20% and 25%, respectively, while thiobarbituric acid reactive substances (TBARS) increases up to five-fold. The increase of haemolysis is negligible suggesting that the erythrocyte membrane is spared at the expense of other sacrificial substrates. While there is a clear relationship between the ozone dose and IL-8 levels, we have noticed that high TAS and PTG values inhibit the cytokine production. This is in line with the current idea that hydrogen peroxide, as a byproduct of O3 decomposition, acts as a messenger for the cytokine induction

Ozone and Ozonated Oils in Skin Diseases: A Review

Although orthodox medicine has provided a variety of topical anti-infective agents, some of them have become scarcely effective owing to antibiotic- and chemotherapeutic-resistant pathogens. For more than a century, ozone has been known to be an excellent disinfectant that nevertheless had to be used with caution for its oxidizing properties. Only during the last decade it has been learned how to tame its great reactivity by precisely dosing its concentration and permanently incorporating the gas into triglycerides where gaseous ozone chemically reacts with unsaturated substrates leading to therapeutically active ozonated derivatives. Today the stability and efficacy of the ozonated oils have been already demonstrated, but owing to a plethora of commercial products, the present paper aims to analyze these derivatives suggesting the strategy to obtain products with the best characteristics

MicroRNA Alterations Induced in Human Skin by Diesel Fumes, Ozone, and UV Radiation

Epigenetic alterations are a driving force of the carcinogenesis process. MicroRNAs play a role in silencing mutated oncogenes, thus defending the cell against the adverse consequences of genotoxic damages induced by environmental pollutants. These processes have been well investigated in lungs; however, although skin is directly exposed to a great variety of environmental pollutants, more research is needed to better understand the effect on cutaneous tissue. Therefore, we investigated microRNA alteration in human skin biopsies exposed to diesel fumes, ozone, and UV light for over 24 h of exposure. UV and ozone-induced microRNA alteration right after exposure, while the peak of their deregulations induced by diesel fumes was reached only at the end of the 24 h. Diesel fumes mainly altered microRNAs involved in the carcinogenesis process, ozone in apoptosis, and UV in DNA repair. Accordingly, each tested pollutant induced a specific pattern of microRNA alteration in skin related to the intrinsic mechanisms activated by the specific pollutant. These alterations, over a short time basis, reflect adaptive events aimed at defending the tissue against damages. Conversely, whenever environmental exposure lasts for a long time, the irreversible alteration of the microRNA machinery results in epigenetic damage contributing to the pathogenesis of inflammation, dysplasia, and cancer induced by environmental pollutants

Lipid-Based Nanosystems for the Topical Application of Ferulic Acid: A Comparative Study

In this study, we examined and compared two different lipid-based nanosystems (LBNs), namely Transferosomes (TFs) and Monoolein Aqueous Dispersions (MADs), as delivery systems for the topical application of Ferulic Acid (FA), an antioxidant molecule derived from natural sources. Our results, as demonstrated through Franz-cell experiments, indicate that the LBNs produced with poloxamer 188 in their composition create a multilamellar system. This system effectively controls the release of the drug. Nonetheless, we found that the type of non-ionic surfactant can impact the drug release rate. Regarding FA diffusion from the MAD, this showed a lower diffusion rate compared with the TF. In terms of an in vivo application, patch tests revealed that all LBN formulations tested were safe when applied under occlusive conditions for 48 h. Ad-ditionally, human skin biopsies were used to determine whether FA-containing formulations could influence skin tissue morphology or provide protection against O3 exposure. Analyses sug-gest that treatment with TFs composed of poloxamer 188 and MAD formulations might protect against structural skin damage (as observed in hematoxylin/eosin staining) and the development of an oxidative environment (as indicated by 4-hyroxinonenal (4HNE) expression levels) induced by O3 exposure. In contrast, formulations without the active ingredient did not offer protection against the detrimental effects of O3 exposur

Altered inflammasome machinery as a key player in the perpetuation of Rett syndrome oxinflammation

Rett syndrome (RTT) is a progressive neurodevelopmental disorder mainly caused by mutations in the X-linked MECP2 gene. RTT patients show multisystem disturbances associated with an oxinflammatory status. Inflammasomes are multi-protein complexes, responsible for host immune responses against pathogen infections and redox-related cellular stress. Assembly of NLRP3/ASC inflammasome triggers pro-caspase-1 activation, thus, resulting in IL-1β and IL-18 maturation. However, an aberrant activation of inflammasome system has been implicated in several human diseases. Our aim was to investigate the possible role of inflammasome in the chronic subclinical inflammatory condition typical of RTT, by analyzing this complex in basal and lipopolysaccharide (LPS)+ATP-stimulated primary fibroblasts, as well as in serum from RTT patients and healthy volunteers. RTT cells showed increased levels of nuclear p65 and ASC proteins, pro-IL-1β mRNA, and NLRP3/ASC interaction in basal condition, without any further response upon the LPS + ATP stimuli. Moreover, augmented levels of circulating ASC and IL-18 proteins were found in serum of RTT patients, which are likely able to amplify the inflammatory response. Taken together, our findings suggest that RTT patients exhibited a challenged inflammasome machinery at cellular and systemic level, which may contribute to the subclinical inflammatory state feedback observed in this pathology

Cigarette Smoke Affects Keratinocytes SRB1 Expression and Localization via H2O2 Production and HNE Protein Adducts Formation

Scavenger Receptor B1 (SR-B1), also known as HDL receptor, is involved in cellular cholesterol uptake. Stratum corneum (SC), the outermost layer of the skin, is composed of more than 25% cholesterol. Several reports support the view that alteration of SC lipid composition may be the cause of impaired barrier function which gives rise to several skin diseases. For this reason the regulation of the genes involved in cholesterol uptake is of extreme significance for skin health. Being the first shield against external insults, the skin is exposed to several noxious substances and among these is cigarette smoke (CS), which has been recently associated with various skin pathologies. In this study we first have shown the presence of SR-B1 in murine and human skin tissue and then by using immunoblotting, immunoprecipitation, RT-PCR, and confocal microscopy we have demonstrated the translocation and the subsequent lost of SR-B1 in human keratinocytes (cell culture model) after CS exposure is driven by hydrogen peroxide (H2O2) that derives not only from the CS gas phase but mainly from the activation of cellular NADPH oxidase (NOX). This effect was reversed when the cells were pretreated with NOX inhibitors or catalase. Furthermore, CS caused the formation of SR-B1-aldheydes adducts (acrolein and 4-hydroxy-2-nonenal) and the increase of its ubiquitination, which could be one of the causes of SR-B1 loss. In conclusion, exposure to CS, through the production of H2O2, induced post-translational modifications of SR-B1 with the consequence lost of the receptor and this may contribute to the skin physiology alteration as a consequence of the variation of cholesterol uptake

HelixComplex snail mucus as a potential technology against O3 induced skin damage

Mucus form H. aspersa muller has been reported to have several therapeutic proprieties, such as antimicrobial activity, skin protection and wound repair. In this study, we have analyzed H. aspersa mucus (Helixcomplex) bio-adhesive efficacy and its defensive properties against the ozone (O3) (0.5 ppm for 2 hours) exposure in human keratinocytes and reconstructed human epidermis models. Cytotoxicity, tissue morphology and cytokine levels were determined. We confirmed HelixComplex regenerative and bio-adhesive properties, the latter possibly via the characteristic mucopolysaccharide composition. In addition, HelixComplex was able to protect from O3 exposure by preventing oxidative damage and the consequent pro-inflammatory response in both 2D and 3D models. Based on this study, it is possible to suggest HelixComplex as a potentially new protective technology against pollution induced skin damage

Synthesis and biological evaluation of fluorinated 1,5-diarylpyrrole-3-alkoxyethyl ether derivatives as selective COX-2 inhibitors endowed with anti-inflammatory activity

A series of substituted 1,5-diarylpyrrole-3-alkoxyethyl ethers were previously synthesized and the potential anti-inflammatory and antinociceptive activities of these compounds were evaluated in vivo. The compounds displayed a very good activity against both carrageenan-induced hyperalgesia and oedema in the rat paw test. Therefore, in a very preliminary test, compounds (8a,b) showed antiproliferative activity in the HaCaT (aneuploid immortal keratinocyte from adult human skin) cell models. On these basis, our research continued with the synthesis of fluorinated derivatives (8c,d, 9b-d, and 10b-d) with the aim of improving the pharmacokinetic profile of the previous active compounds. Substitution of a hydrogen atom by a fluorine atom may change the conformational preferences of the molecules due to stereoelectronic effects and also fluorine atom may be able to exert the metabolic obstruction reducing the "first-pass effect". Compound 10b exhibited a prominent in vivo anti-inflammatory and antinociceptive activities, in addition its antiproliferative power in an in vitro model of human skin cancer is herein described