Evolution and regulation of photoprotective mechanisms in microalgae

La fotosintesi ossigenica \ue8 un processo per mezzo del quale l\u2019energia solare e l\u2019anidride carbonica (CO2) sono utilizzate per produrre ossigeno (O2) e biomassa. La conversione dell\u2019energia luminosa in energia chimica \ue8 condotta da complessi multiproteici denominati Fotosistema II (PSII) e Fotosistema I (PSI). Il PSII e il PSI mediano la separazione di carica, la raccolta della luce e il trasporto di elettroni dall\u2019acqua, producendo il potere riducente necessario per fissare la CO2 in carboidrati (ATP e NADPH). I Fotosistemi sono composti da due unit\ue0 principali: il centro di reazione, sito in cui avvengono le reazioni biochimiche e la separazione di carica, e il sistema antenna, costituito da complessi proteici di raccolta della luce (LHC), coinvolti principalmente nella raccolta della luce e nel trasferimento dell\u2019energia d\u2019eccitazione al centro di reazione. Gli organismi fotosintetici sfruttano la radiazione fotosinteticamente attiva (PAR) a fini metabolici. Variazioni nell'irradianza, quali l\u2019eccesso di luce, possono determinare condizioni limitanti o di stress, portando alla formazione di specie reattive dell\u2019ossigeno (ROS), le quali, influenzando la crescita delle piante e ne riducono la produttivit\ue0. L'attivazione del processo di dissipazione termica, denominato Non-Photochemical Quenching (NPQ), ha un ruolo fondamentale nella reazione di quenching (smorzamento) degli stati eccitati di singoletto della clorofilla, dissipando l'energia di eccitazione sotto forma di calore, prevenendo quindi lo stress foto-ossidativo. Nelle microalghe fino all'80% dell'energia luminosa assorbita pu\uf2 essere riemessa sotto forma di calore con conseguente riduzione della produttivit\ue0 totale di biomassa. Questa tesi \ue8 incentrata sullo studio della regolazione dell\u2019NPQ in diverse specie di alghe. A tale scopo sono stati applicati diversi approcci quali la trasformazione genetica, la caratterizzazione fenotipica e spettroscopica di cellule intere e di complessi proteici isolati. La regolazione dell\u2019NPQ a livello del PSII e del PSI \ue8 stata ampiamente studiata anche in relazione alle proteine LHC nell\u2019organismo modello per le alghe verdi, Chlamydomonas reinhardtii, in cui \ue8 stata evidenziata un'attivit\ue0 di quenching in entrambi i Fotosistemi. Nella specie di microalga di uso commerciale, Chlorella vulgaris, la regolazione dell\u2019NPQ \ue8 stata studiata in relazione all'accumulo di zeaxantina, evidenziandone una forte dipendenza. Infine, la regolazione fotosintetica \ue8 stata monitorata in cellule intere e in complessi isolati nella microalga Haematococcus pluvialis, in presenza di forti stress indotti dall\u2019eccesso di energia luminosa e dalla carenza di nutrienti.Oxygenic photosynthesis is a process by which sunlight energy and CO2 are used to produce O2 and biomass. The light energy conversion into chemical energy is carried forth by multiproteic complexes called Photosystem II (PSII) and Photosystem I (PSI). PSII and PSI drive charge separation, light harvesting and electron transport from water, producing the reducing power necessary to fix CO2 into carbohydrates (ATP and NADPH). Photosystems are composed by two moieties: a core reaction center, site of biochemical reactions and charge separation, and an antenna system constituted by Light Harvesting Complex (LHC) proteins mainly involved in light harvesting and excitation energy transfer to the reaction center. Photosynthetic organisms use the photosynthetically active radiation (PAR) for their metabolic processes but irradiance undergo changes and light excess becomes a limit or even a stressor leading to the formation of Reactive Oxygen Species (ROS) which influence plant growth and could decrease crop productivity. Photo-oxidative stress can be prevented by activation of thermal dissipation process called Non-Photochemical Quenching (NPQ), which has an important role in quenching chlorophylls singlet excited states dissipating the excitation energy in form of heat. In microalgae, up to 80% of absorbed light energy can be re-emitted as heat with a consequent reduction of total biomass productivity. This thesis was focused into the investigation of NPQ regulation in several algae species. For this purpose, different approaches were applied including genetic transformation, phenotypic and spectroscopic characterization of entire cells and of isolated complexes. The NPQ regulation at the level of both PSII and PSI also in relation with LHC proteins was fully investigated in the model green alga Chlamydomonas reinhardtii evidencing a quenching activity in both Photosystems. In the commercial microalga specie, Chlorella vulgaris, the NPQ regulation was studied in relation with zeaxanthin accumulation evidencing a strong dependency. Finally, in the microalga Haematococcus pluvialis, the photosynthetic regulation was also monitored in entire cells and isolated complexes, in presence of strong stresses induced by light excess and nutrients depletion

Functional analysis of photosynthetic pigment binding complexes in the green alga Haematococcus pluvialis reveals distribution of astaxanthin in Photosystems

Astaxanthin is a ketocarotenoid produced by photosynthetic microalgae. It is a pigment of high industrial interest in acquaculture, cosmetics, and nutraceutics due to its strong antioxidant power. Haematococcus pluvialis, a fresh-water microalga, accumulates high levels of astaxanthin upon oxidative stress, reaching values up to 5% per dry weight. H. pluvialis accumulates astaxanthin in oil droplets in the cytoplasm, while the chloroplast volume is reduced. In this work, we investigate the biochemical and spectroscopic properties of the H. pluvialis pigment binding complexes responsible for light harvesting and energy conversion. Our findings demonstrate that the main features of chlorophyll and carotenoid binding complexes previously reported for higher plants or Chlamydomonas reinhardtii are preserved under control conditions. Transition to astaxanthin rich cysts however leads to destabilization of the Photosystems. Surprisingly, astaxanthin was found to be bound to both Photosystem I and II, partially substituting β-carotene, and thus demonstrating possible astaxanthin biosynthesis in the plastids or transport from the cytoplasm to the chloroplast. Astaxanthin binding to Photosystems does not however improve their photoprotection, but rather reduces the efficiency of excitation energy transfer to the reaction centers. We thus propose that astaxanthin binding partially destabilizes Photosystem I and II

Validation of epidemiological tools for eczema diagnosis in brazilian children: the isaac's and uk working party's criteria

BACKGROUND: Instruments for field diagnosis of eczema are increasingly used, and it is essential to understand specific limitations to make best use of their strengths. Our objective was to assess the validity of ISAAC and UK Working Party criteria for field diagnosis of eczema in children. METHODS: We performed a cohort study in urban Brazil. Parents/guardians of 1,419 children answered ISAAC phase II questionnaire. Children were examined for skin lesions (UKWP protocol). Two dermatologists examined most cases of eczema (according to ISAAC or UKWP), and a sample without eczema. RESULTS: Agreement between repeat questionnaires on the filter question was poor (kappa = 0.4). Agreement between the 2 dermatologists was fair (kappa = 0.6). False positive reports included scabies in 39% of ISAAC cases and 33% of UKWP cases. Sensitivity and PPV were low (ISAAC: 37.1% and 16.1%; UKWP: 28.6% and 23.8%). Specificity and NPV were high (ISAAC: 90.0% and 96.6%; UKWP: 95.3% and 96.2%). One-year prevalence of eczema was 11.3% (ISAAC), 5.9% (UKWP) and 4.9% (adjusted dermatologist diagnosis). Point prevalence of scabies (alone or not) was 43%, 33% and 18%, in eczemas according to ISAAC, to UKWP and to dermatologists. The reasons why children with eczema were not identified by ISAAC or UKWP were wrongly denying dry skin, itchy rash or personal history of atopic diseases. A limitation is that questionnaire was already validated in Brazil, but not field tested in this specific setting. CONCLUSIONS: Studies using UKWP or ISAAC criteria should include a validation arm, to contribute to the understanding of potential limitations of their use in different contexts and to explore solutions. We list specific recommendations

Long-Term Drug Survival and Effectiveness of Secukinumab in Patients with Moderate to Severe Chronic Plaque Psoriasis: 42-Month Results from the SUPREME 2.0 Study

Purpose: SUPREME, a phase IIIb study conducted in Italy, demonstrated safety and high efficacy of secukinumab for up to 72 weeks in patients with moderate-to-severe plaque-type psoriasis. SUPREME 2.0 study aimed to provide real-world data on the long-term drug survival and effectiveness of secukinumab beyond 72 weeks. Patients and Methods: SUPREME 2.0 is a retrospective observational chart review study conducted in patients previously enrolled in SUPREME study. After the end of the SUPREME study, eligible patients continued treatment as per clinical practice, and their effectiveness and drug survival data were retrieved from medical charts. Results: Of the 415 patients enrolled in the SUPREME study, 297 were included in SUPREME 2.0; of which, 210 (70.7%) continued secukinumab treatment throughout the 42-month observation period. Patients in the biologic-naïve cohort had higher drug survival than those in the biologic-experienced cohort (74.9% vs 61.7%), while HLA-Cw6–positive and HLA-Cw6–negative patients showed similar drug survival (69.3% and 71.9%). After 42 months, Psoriasis Area and Severity Index (PASI) 90 was achieved by 79.6% of patients overall; with a similar proportion of biologic-naïve and biologic-experienced patients achieving PASI90 (79.8% and 79.1%). The mean absolute PASI score reduced from 21.94 to 1.38 in the overall population, 21.90 to 1.24 in biologic-naïve and 22.03 to 1.77 in biologic-experienced patients after 42 months. The decrease in the absolute PASI score was comparable between HLACw6–positive and HLA–Cw6-negative patients. The baseline Dermatology Life Quality Index scores also decreased in the overall patients (10.5 to 2.32) and across all study sub-groups after 42 months. Safety was consistent with the known profile of secukinumab, with no new findings. Conclusion: In this real-world cohort study, secukinumab showed consistently high long-term drug survival and effectiveness with a favourable safety profile

Long-Term Drug Survival and Effectiveness of Secukinumab in Patients with Moderate to Severe Chronic Plaque Psoriasis: 42-Month Results from the SUPREME 2.0 Study

Purpose: SUPREME, a phase IIIb study conducted in Italy, demonstrated safety and high efficacy of secukinumab for up to 72 weeks in patients with moderate-to-severe plaque-type psoriasis. SUPREME 2.0 study aimed to provide real-world data on the long-term drug survival and effectiveness of secukinumab beyond 72 weeks.Patients and Methods: SUPREME 2.0 is a retrospective observational chart review study conducted in patients previously enrolled in SUPREME study. After the end of the SUPREME study, eligible patients continued treatment as per clinical practice, and their effectiveness and drug survival data were retrieved from medical charts.Results: Of the 415 patients enrolled in the SUPREME study, 297 were included in SUPREME 2.0; of which, 210 (70.7%) continued secukinumab treatment throughout the 42-month observation period. Patients in the biologic-naive cohort had higher drug survival than those in the biologic-experienced cohort (74.9% vs 61.7%), while HLA-Cw6-positive and HLA-Cw6-negative patients showed similar drug survival (69.3% and 71.9%). After 42 months, Psoriasis Area and Severity Index (PASI) 90 was achieved by 79.6% of patients overall; with a similar proportion of biologic-naive and biologic-experienced patients achieving PASI90 (79.8% and 79.1%). The mean absolute PASI score reduced from 21.94 to 1.38 in the overall population, 21.90 to 1.24 in biologic-naive and 22.03 to 1.77 in biologic-experienced patients after 42 months. The decrease in the absolute PASI score was comparable between HLA-Cw6-positive and HLA-Cw6-negative patients. The baseline Dermatology Life Quality Index scores also decreased in the overall patients (10.5 to 2.32) and across all study sub-groups after 42 months. Safety was consistent with the known profile of secukinumab, with no new findings. Conclusion: In this real-world cohort study, secukinumab showed consistently high long-term drug survival and effectiveness with a favourable safety profile

Activin A Induces Langerhans Cell Differentiation In Vitro and in Human Skin Explants

Langerhans cells (LC) represent a well characterized subset of dendritic cells located in the epidermis of skin and mucosae. In vivo, they originate from resident and blood-borne precursors in the presence of keratinocyte-derived TGFβ. Ιn vitro, LC can be generated from monocytes in the presence of GM-CSF, IL-4 and TGFβ. However, the signals that induce LC during an inflammatory reaction are not fully investigated. Here we report that Activin A, a TGFβ family member induced by pro-inflammatory cytokines and involved in skin morphogenesis and wound healing, induces the differentiation of human monocytes into LC in the absence of TGFβ. Activin A-induced LC are Langerin+, Birbeck granules+, E-cadherin+, CLA+ and CCR6+ and possess typical APC functions. In human skin explants, intradermal injection of Activin A increased the number of CD1a+ and Langerin+ cells in both the epidermis and dermis by promoting the differentiation of resident precursor cells. High levels of Activin A were present in the upper epidermal layers and in the dermis of Lichen Planus biopsies in association with a marked infiltration of CD1a+ and Langerin+ cells. This study reports that Activin A induces the differentiation of circulating CD14+ cells into LC. Since Activin A is abundantly produced during inflammatory conditions which are also characterized by increased numbers of LC, we propose that this cytokine represents a new pathway, alternative to TGFβ, responsible for LC differentiation during inflammatory/autoimmune conditions

The aetiology of eyelid dermatitis in a series of 191 patients

Several studies identified allergiccontact dermatitis (ACD) as the most commoncause of periorbital dermatitis. Nevertheless,there is paucity of studies directly comparing therelevance of allergens in the eyelid dermatitis versusnon eyelid dermatitis populations.Methods. 1043 patients who presented with the clinicalsuspicion of ACD were patch tested in a 2-yearperiod. Patient demographics, clinical features, historyof atopy and results of patch tests were collected.Patch test results of patients with eyelid dermatitiswere compared with those of patients withdermatitis confined to other body areas.Results. 191 out of 1043 patch tested patients(18.3%) presented with eyelid dermatitis. Patientswith eyelid dermatitis were more likely to bewomen compared to the non-eyelid group ofpatients. The two groups did not differ as for thefrequency of atopy, including respiratory atopyand atopic dermatitis. The prevalence of ACD wasstatistically higher in the eyelid group (64.4%)compared to the non-eyelid group (49.4%). Themost relevant allergens in patients with eyelid dermatitiswere metals, preservatives and fragrances.Allergens resulting statistically more frequent ineyelid patients were methylchloroisothiazolinone/methylisothiazolinone, nickel sulphate, disperseblue 124, dimethylaminopropylamine.Conclusions. Patients with eyelid dermatitis arelikely to have ACD, and need appropriate patchtesting

Psychological Stress and Salivary Cortisol Levels in Patients with Plaque Psoriasis

Psychological stress has long been recognized as a trigger for plaque psoriasis, and pre- liminary evidence suggests that psoriasis could be associated with alterations in the hypothalamic- pituitary-adrenal (HPA) axis, resulting in impaired cortisol response to stress. This study aimed to investigate psychological stress, anxiety, depression and salivary cortisol in psoriatic patients. A cross sectional study involving 126 adult patients with plaque psoriasis and 116 adult healthy controls was conducted. Demographic, clinical data, Perceived Stress Scale (PSS) and Hospital Anxiety and Depression Scale (HADS) were collected. Cases and controls were asked whether they felt stressed in the last month, whilst psoriatic patients were also interrogated whether they found that psoriasis could have been worsened by stress. Moreover, 54 randomly selected subjects (27 psoriasis patients and 27 controls) underwent salivary cortisol testing at 8 am. PSS, HADS depression and anxiety subscales were significantly higher in psoriatic patients than in controls (17.2 ± 0.6 vs. 15.1 ± 0.8 p = 0.0289), (9.5 ± 0.3 vs. 6.2 ± 0.3 p < 0.001) and (8.2 ± 0.4 vs. 4.2 ± 0.3 p < 0.001), respectively. A higher rate of psoriatic patients reported feeling stress over the last month (45% vs. 19%, p < 0.001), and stress was considered a potential trigger for psoriasis flare-ups in 69% of cases. Psoriasis was strongly associated with higher PSS and HADS scores independently of sex, body mass index, dia- betes, hypertension, dyslipidemia, and occupational status. Salivary cortisol was significantly lower in psoriatic patients compared to controls (9.6 ± 0.5 vs. 14.0 ± 1.1 nmol/L, p < 0.001). In conclusion, psoriasis was associated with higher psychological stress, anxiety and depressive symptoms, and with impaired cortisol response to stress

Overview of Atopic Dermatitis in Different Ethnic Groups

Atopic dermatitis (AD) is a common chronic inflammatory skin disease with a high prevalence worldwide, including countries from Asia, Africa, and Latin America, and in different ethnic groups. In recent years, more attention has been placed on the heterogeneity of AD associated with multiple factors, including a patient’s ethnic background, resulting in an increasing body of clinical, genetic, epidemiologic, and immune-phenotypic evidence that delineates differences in AD among racial groups. Filaggrin (FLG) mutations, the strongest genetic risk factor for the development of AD, are detected in up to 50% of European and 27% of Asian AD patients, but very rarely in Africans. Th2 hyperactivation is a common attribute of all ethnic groups, though the Asian endotype of AD is also characterized by an increased Th17-mediated signal, whereas African Americans show a strong Th2/Th22 signature and an absence of Th1/Th17 skewing. In addition, the ethnic heterogeneity of AD may hold important therapeutic implications as a patient’s genetic predisposition may affect treatment response and, thereby, a tailored strategy that better targets the dominant immunologic pathways in each ethnic subgroup may be envisaged. Nevertheless, white patients with AD represent the largest ethnicity enrolled and tested in clinical trials and the most treated in a real-world setting, limiting investigations about safety and efficacy across different ethnicities. The purpose of this review is to describe the heterogeneity in the pathophysiology of AD across ethnicities and its potential therapeutic implications

Photosynthetic response to nitrogen starvation and high light in Haematococcus pluvialis

Astaxanthin is a carotenoid mainly produced by microalgae upon exposure to stress conditions: this pigment has anti-oxidant, anti-inflammatory and anti-cancer capacity and it is widely used as pigmentation agent in different industrial sectors. Abiotic stresses such as exposure to high irradiances and/or nitrogen starvation are commonly used to induce astaxanthin biosynthesis in freshwater green alga Haematococcus pluvialis. In this work high light and nitrogen deprivation were applied as single or combined stresses in order to investigate their influence on the photosynthetic properties of H. pluvialis cultures. The results reported here demonstrate that nitrogen starvation inhibits chlorophyll biosynthesis and favors chlorophyll b degradation, chlororespiration and cyclic electron transport, while cells grown in high light are characterized by a higher destabilization of PSII. The combination of high light and nitrogen deprivation induced the highest astaxanthin production and also the fastest photoprotective response which cooperatively prevented Photosystem II from the damage observed in high light stress and nitrogen supplemented medium. In these conditions inhibition of astaxanthin accumulation leads to a reduced cell size but does not induce a higher photosensitivity of photosynthetic machinery