Rebeca Pérez

Amplification Mechanisms of Inflammation: Paracrine Stimulation of Arachidonic Acid Mobilization by Secreted Phospholipase A2 Is Regulated by Cytosolic Phospholipase A2-Derived Hydroperoxyeicosatetraenoic Acid

In macrophages and other major immunoinflammatory cells, two phospholipase A2 (PLA2) enzymes act in concert to mobilize arachidonic acid (AA) for immediate PG synthesis, namely group IV cytosolic phospholipase A2 (cPLA2) and a secreted phospholipase A2 (sPLA2). In this study, the molecular mechanism underlying cross-talk between the two PLA2s during paracrine signaling has been investigated. U937 macrophage-like cells respond to Con A by releasing AA in a cPLA2-dependent manner, and addition of exogenous group V sPLA2 to the activated cells increases the release. This sPLA2 effect is abolished if the cells are pretreated with cPLA2 inhibitors, but is restored by adding exogenous free AA. Inhibitors of cyclooxygenase and 5-lipoxygenase have no effect on the response to sPLA2. In contrast, ebselen strongly blocks it. Reconstitution experiments conducted in pyrrophenone-treated cells to abolish cPLA2 activity reveal that 12- and 15-hydroperoxyeicosatetraenoic acid (HPETE) are able to restore the sPLA2 response to levels found in cells displaying normal cPLA2 activity. Moreover, 12- and 15-HPETE are able to enhance sPLA2 activity in vitro, using a natural membrane assay. Neither of these effects is mimicked by 12- or 15-hydroxyeicosatetraenoic acid, indicating that the hydroperoxy group of HPETE is responsible for its biological activity. Collectively, these results establish a role for 12/15-HPETE as an endogenous activator of sPLA2-mediated phospholipolysis during paracrine stimulation of macrophages and identify the mechanism that connects sPLA2 with cPLA2 for a full AA mobilization response.

Involvement of Group VIA Calcium-Independent Phospholipase A2 in Macrophage Engulfment of Hydrogen Peroxide-Treated U937 Cells

Hydrogen peroxide-induced apoptosis of U937 cells results in substantial hydrolysis of membrane phospholipids by calcium-independent group VIA phospholipase A2 (iPLA2-VIA). However, abrogation of cellular iPLA2-VIA neither delays nor decreases apoptosis, suggesting that, beyond a mere destructive role, iPLA2-VIA may serve other specific roles. In this study, we report that phagocytosis of apoptosing U937 cells by macrophages is blunted if the cells are depleted of iPLA2-VIA by treatment with an inhibitor or an antisense oligonucleotide, and it is augmented by overexpression of iPLA2-VIA in the dying cells. Thus, the magnitude of macrophage phagocytosis correlates with the level of iPLA2-VIA activity of the dying cells. Eliminating by antisense oligonucleotide technology of cytosolic group IVA phospholipase A2 does not attenuate phagocytosis of U937 dying cells by macrophages. Incubation of U937 cells with different fatty acids has no effect on either the extent of hydrogen peroxide-induced apoptosis or the degree of phagocytosis of the dying cells by macrophages. However, preincubation of the macrophages with lysophosphatidylcholine before exposing them to the dying cells blocks phagocytosis of the latter. These results indicate that formation of lysophosphatidylcholine by iPLA2-VIA in hydrogen peroxide-treated U937 cells to induce apoptosis directly contributes to their efficient clearance by macrophages.

Evaluation of wastewater treatment in a novel anoxic-aerobic algal-bacterial photobioreactor with biomass recycling through carbon and nitrogen mass balances.

Algal-bacterial symbiosis, implemented in an innovative anoxic-aerobic photobioreactor configuration with biomass recycling, supported an efficient removal of total organic carbon (86-90%), inorganic carbon (57-98%) and total nitrogen (68-79%) during synthetic wastewater treatment at a hydraulic and sludge retention times of 2 days and 20 days, respectively. The availability of inorganic carbon in the photobioreactor, determined by its supply in the wastewater and microalgae activity, governed the extent of nitrogen removal by assimilation or nitrification-denitrification. Unexpectedly, nitrate production was negligible despite the high dissolved oxygen concentrations, denitrification being only based on nitrite reduction. Biomass recycling resulted in the enrichment of rapidly settling algal flocs, which supported effluent total suspended solid concentrations below the European Union maximum discharge limits. Finally, the maximum nitrous oxide emissions recorded were far below the emission factors reported for wastewater treatment plants, confirming the environmental sustainability of this innovative photobioreactor in terms of global warming impact.

Exploring the potential of fungi for methane abatement: Performance evaluation of a fungal-bacterial biofilter.

Despite several fungal strains have been retrieved from methane-containing environments, the actual capacity and role of fungi on methane abatement is still unclear. The batch biodegradation tests here performed demonstrated the capacity of Graphium sp. to co-metabolically biodegrade methane and methanol. Moreover, the performance and microbiology of a fungal-bacterial compost biofilter treating methane at concentrations of ∼2% was evaluated at empty bed residence times of 40 and 20 min under different irrigation rates. The daily addition of 200 mL of mineral medium resulted in elimination capacities of 36.6 ± 0.7 g m(-3) h(-1) and removal efficiencies of ≈90% at the lowest residence time. The indigenous fungal community of the compost was predominant in the final microbial population and outcompeted the inoculated Graphium sp. during biofilter operation.

Microaerobic desulphurisation unit: A new biological system for the removal of H2S from biogas

A new biotechnology for the removal of H2S from biogas was devised. The desulphurisation conditions present in microaerobic digesters were reproduced inside an external chamber called a microaerobic desulphurisation unit (MDU). A 10 L-unit was inoculated with 1L of digested sludge in order to treat the biogas produced in a pilot digester. During the 128 d of research under such conditions, the average removal efficiency was 94%. The MDU proved to be robust against fluctuations in biogas residence time (57-107 min), inlet H2S concentration (0.17-0.39% v/v), O2/H2S supplied ratio (17.3-1.4 v/v), and temperature (20-35°C). Microbiological analysis confirmed the presence of at least three genera of sulphide-oxidising bacteria. Approximately 60% of all the H2S oxidised was recovered from the bottom of the system in the form of large solid S(0) sheets with 98% w/w of purity. Therefore, this system could become a cost-effective alternative to the conventional biotechniques for biogas desulphurisation.

Microaerobic digestion of sewage sludge on an industrial-pilot scale: the efficiency of biogas desulphurisation under different configurations and the impact of O2 on the microbial communities.

Biogas produced in an industrial-pilot scale sewage sludge reactor (5m(3)) was desulphurised by imposing microaerobic conditions. The H2S concentration removal efficiency was evaluated under various configurations: different mixing methods and O2 injection points. Biogas was entirely desulphurised under all the configurations set, while the O2 demand of the digester decreased over time. Although the H2S removal seemed to occur in the headspace, S(0) (which was found to be the main oxidation product) was scarcely deposited there in the headspace. O2 did not have a significant impact on the digestion performance; the VS removal remained around 47%. Conversely, DGGE revealed that the higher O2 transfer rate to the sludge maintained by biogas recirculation increased the microbial richness and evenness, and caused an important shift in the structure of the bacterial and the archaeal communities in the long term. All the archaeal genera identified (Methanosaeta, Methanospirillum and Methanoculleus) were present under both anaerobic and microaerobic conditions.

The headspace of microaerobic reactors: sulphide-oxidising population and the impact of cleaning on the efficiency of biogas desulphurisation.

O2-limiting/microaerobic conditions were applied in order to control the H2S content of biogas. The S(0)-rich deposits found all over the headspace of two pilot reactors (R1 and R2) as a result of operating under such conditions for 7 and 15 months (respectively) were sampled and removed. After restarting micro-oxygenation, H2S-free biogas was rapidly obtained, and the O2 demand of R2 decreased. This highlighted the need for a cleaning interval of less than 14 months in order to minimise the micro-oxygenation cost. The H2S removed from R2 after approximately 1 month was recovered from its headspace as S(0), thus indicating that the biogas desulphurisation did not take place at the liquid interface. Denaturing gradient gel electrophoresis indicated that the composition, species richness and size of the sulphide-oxidising bacteria population depended on the location, and, more specifically, moisture availability, and indicated increasing species richness over time. Additionally, a possible succession was estimated.

Two-liquid phase partitioning biotrickling filters for methane abatement: exploring the potential of hydrophobic methanotrophs.

The potential of two-liquid phase biotrickling filters (BTFs) to overcome mass transfer limitations derived from the poor aqueous solubility of CH4 has been scarcely investigated to date. In this context, the abatement of diluted methane emissions in two-liquid phase BTFs was evaluated using two different inocula: a type II methanotrophs culture in BTF 1 and a hydrophobic microbial consortium capable of growing inside silicone oil in BTF 2. Both BTFs supported stable elimination capacities above 45 g m(-3) h(-1) regardless of the inoculum, whereas no improvement derived from the presence of hydrophobic microorganisms compared to the type II metanotrophs culture was observed. Interestingly, the addition of silicone oil mediated a reduced metabolites concentration in the recycling aqueous phase, thus decreasing the needs for mineral medium renewal. Moreover, a 78% similarity was recorded between the microbial communities enriched in both BTFs at the end of the experimental period in spite of the differences in the initial inoculum structure. The results obtained confirmed the superior performance of two-liquid phase BTFs for CH4 abatement compared with conventional biotrickling filters.

Biological anoxic treatment of O2-free VOC emissions from the petrochemical industry: A proof of concept study

An innovative biofiltration technology based on anoxic biodegradation was proposed in this work for the treatment of inert VOC-laden emissions from the petrochemical industry. Anoxic biofiltration does not require conventional O2 supply to mineralize VOCs, which increases process safety and allows for the reuse of the residual gas for inertization purposes in plant. The potential of this technology was evaluated in a biotrickling filter using toluene as a model VOC at loads of 3, 5, 12 and 34 g m(-3)h(-1) (corresponding to empty bed residence times of 16, 8, 4 and 1.3 min) with a maximum elimination capacity of ∼3 g m(-3)h(-1). However, significant differences in the nature and number of metabolites accumulated at each toluene load tested were observed, o- and p-cresol being detected only at 34 g m(-3)h(-1), while benzyl alcohol, benzaldehyde and phenol were detected at lower loads. A complete toluene removal was maintained after increasing the inlet toluene concentration from 0.5 to 1 g m(-3) (which entailed a loading rate increase from 3 to 6 g m(-3)h(-1)), indicating that the system was limited by mass transfer rather than by biological activity. A high bacterial diversity was observed, the predominant phyla being Actinobacteria and Proteobacteria.

Anoxic biodegradation of BTEX in a biotrickling filter

Emissions of BTEX (benzene, toluene, ethylbenzene and xylene) from the petrochemical industry are characterized by a low pollutants concentration and the absence of oxygen. Biodegradation of these pollutants using nitrate as the electron acceptor is of key interest to reuse the residual gas for inertization purposes. However, the biological mineralization of BTEX is often limited by their recalcitrant nature and the toxicity of the secondary metabolites produced. The potential of an anoxic biotrickling filter for the treatment of a model O2-free BTEX-laden emission at inlet individual concentrations of ~700mgm-3 was here evaluated. A UV oxidation step was also tested both in the recycling liquid and in the inlet gas emission prior to biofiltration. Removal efficiencies >90% were achieved for both toluene and ethylbenzene, corresponding to elimination capacities (ECs) of 1.4±0.2gm-3h-1 and 1.5±0.3gm-3h-1, respectively, while ~45% of xylene (EC=0.6±0.1g m-3h-1) was removed at a liquid recycling rate of 2mh-1. Benzene biodegradation was however limited by the accumulation of toxic metabolites in the liquid phase. The oxidation of these intermediates in the recycling liquid by UV photolysis boosted benzene abatement, achieving an average EC of 0.5±0.2gm-3h-1 and removals of ~40%. However, the implementation of UV oxidation as a pretreatment step in the inlet gas emission resulted in the deterioration of the BTEX biodegradation capacity of the biotrickling filter. Finally, a high bacterial diversity was observed throughout the entire experiment, the predominant phyla being Proteobacteria and Deinococcus-thermus.