Browsing by Subject "FLUORESCENCE"

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  • Kortesoja, Maarit; Trofin, Raluca Elena; Hanski, Leena (2020)
    The obligate intracellular bacterium, Chlamydia pneumoniae, has been identified as a risk factor for several chronic inflammatory diseases in addition to respiratory tract infections. The dissemination of C. pneumoniae from respiratory tract to secondary sites of infection occurs via infected monocyte / macrophage line cells, in which C. pneumoniae can persist as an antibiotic-refractory phenotype. To allow more detailed studies on the epithelium-monocyte/macrophage transition of the infection, new in vitro bioassays are needed. To this end, a coculture system with human continuous cell lines was established. Respiratory epithelial HL cells were infected with C. pneumoniae and THP-1 monocytes were added into the cultures at 67 h post infection. After a 5 h coculture, THP-1 cells were collected with a biotinylated HLA antibody and streptavidin-coated magnetic beads and C. pneumoniae genome copy numbers in THP-1 determined by quantitative PCR. The assay was optimized for cell densities, incubation time, THP-1 separation technique and buffer composition, and its robustness was demonstrated by a Z' value of 0.6. The mitogen-activated protein kinase (MAPK) inhibitors: SP600125 (JNK inhibitor), SB203580 (p38 inhibitor) and FR180204 (ERK inhibitor) suppressed the transfer of C. pneumoniae from HL to THP-1 cells, making them suitable positive controls for the assay. Based on analysis of separate steps of the process, the MAPK inhibitors suppress the bacterial entry to THP-1 cells. The transfer of C. pneumoniae from epithelium to phagocytes represents a crucial step in the establishment of persistent infections by this pathogen, and the presented methods enables future studies to block this process by therapeutic means.
  • Rautaniemi, Kaisa; Zini, Jacopo; Löfman, Emilia; Saari, Heikki; Haapalehto, Iida; Laukka, Johanna; Vesamäki, Sami; Efimov, Alexander; Yliperttula, Marjo; Laaksonen, Timo; Vuorimaa-Laukkanen, Elina; Lisitsyna, Ekaterina S. (2022)
    Studies of extracellular vesicles (EVs), their trafficking and characterization often employ fluorescent labelling. Unfortunately, little attention has been paid thus far to a thorough evaluation of the purification of EVs after labelling, although the presence of an unbound dye may severely compromise the results or even lead to wrong conclusions on EV functionality. Here, we systematically studied five dyes for passive EV labelling and meticulously compared five typical purification methods: ultracentrifugation (UC), ultracentrifugation with discontinuous density gradient (UCG), ultrafiltration (UF), size exclusion chromatography (SEC), and anion exchange chromatography (AEC). A general methodology for evaluation of EV purification efficiency after the labelling was developed and tested to select the purification methods for the chosen dyes. Firstly, we found that some methods initially lead to high EV losses even in the absence of the dye. Secondly, the suitable purification method needs to be found for each particular dye and depends on the physical and chemical properties of the dye. Thirdly, we demonstrated that the developed parameter E-rp (relative purification efficiency) is a useful tool for the pre-screening of the suitable dye-purification method combinations. Additionally, it was also shown that the labelled EVs properly purified from the unbound dye may show significantly reduced contrast and visibility in the target application, e.g. in the live cell fluorescence lifetime imaging.
  • Saarinen, Jukka Kalle Samuel; Gütter, Friederike; Lindman, Mervi M; Agopov, Mikael; Fraser-Miller, Sara J.; Scherließ, Regina; Jokitalo, Eija; Almeida Santos, Helder; Peltonen, Leena; Isomäki, Antti; Strachan, Clare J. (2019)
    A wide variety of nanoparticles are playing an increasingly important role in drug delivery. Label-free imaging techniques are especially desirable to follow the cellular uptake and intracellular fate of nanoparticles. The combined correlative use of different techniques, each with unique advantages, facilitates more detailed investigation about such interactions. The synergistic use of correlative coherent anti-Stokes Raman scattering and electron microscopy (C-CARS-EM) imaging offers label-free, chemically-specific, and (sub)-nanometer spatial resolution for studying nanoparticle uptake into cells as demonstrated in the current study. Coherent anti-Stokes Raman scattering (CARS) microscopy offers chemically-specific (sub)micron spatial resolution imaging without fluorescent labels while transmission electron microscopy (TEM) offers (sub)-nanometer scale spatial resolution and thus visualization of precise nanoparticle localization at the sub-cellular level. This proof-of-concept imaging platform with unlabeled drug nanocrystals and macrophage cells revealed good colocalization between the CARS signal and electron dense nanocrystals in TEM images. The correlative TEM images revealed subcellular localization of nanocrystals inside membrane bound vesicles, showing multivesicular body (MVB)-like morphology typical for late endosomes (LEs), endolysosomes, and phagolysosomes. C-CARS-EM imaging has much potential to study the interactions between a wide range of nanoparticles and cells with high precision and confidence.
  • Owen, Michael C.; Kulig, Waldemar; Rog, Tomasz; Vattulainen, Ilpo; Strodel, Birgit (2018)
    In an effort to delineate how cholesterol protects membrane structure under oxidative stress conditions, we monitored the changes to the structure of lipid bilayers comprising 30 mol% cholesterol and an increasing concentration of Class B oxidized 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) glycerophospholipids, namely, 1-palmitoyl-2-(9'-oxo-nonanoyl)-sn-glycero-3-phosphocholine (PoxnoPC), and 1-palmitoyl-2-azelaoyl-sn-glycero-3-phosphocholine (PazePC), using atomistic molecular dynamics simulations. Increasing the content of oxidized phospholipids (oxPLs) from 0 to 60 mol% oxPL resulted in a characteristic reduction in bilayer thickness and increase in area per lipid, thereby increasing the exposure of the membrane hydrophobic region to water. However, cholesterol was observed to help reduce water injury by moving into the bilayer core and forming more hydrogen bonds with the oxPLs. Cholesterol also resists altering its tilt angle, helping to maintain membrane integrity. Water that enters the 1-nm-thick core region remains part of the bulk water on either side of the bilayer, with relatively few water molecules able to traverse through the bilayer. In cholesterol-rich membranes, the bilayer does not form pores at concentrations of 60 mol% oxPL as was shown in previous simulations in the absence of cholesterol.
  • Haraguchi, Lumi; Asmala, Eero; Jakobsen, Hans H.; Carstensen, Jacob (2019)
    Dissolved organic matter (DOM) is an important component of nutrient cycling, but the role of different organisms controlling the processing of autochthonous DOM remains poorly understood. Aiming to characterize phytoplankton-derived DOM and the effects of complex pelagic communities on its dynamics, we incubated natural plankton communities from a temperate mesohaline estuary under controlled conditions for 18 days. The incubations were carried out in contrasting seasons (spring and autumn) and changes in the planktonic community (phytoplankton, bacteria and microzooplankton), nutrients and DOM were assessed. Our results highlight the complexity of DOM production and fate in natural planktonic communities. Small changes in DOM composition were observed in the experiments relative to the orders-of-magnitude variations experienced in the phytoplankton assembly. We argue that the tight coupling between microbial processing and DOM production by phytoplankton and grazers stabilizes variations in quantity and characteristics of autochthonous DOM, resulting in apparently homogeneous semi-labile DOM pool throughout the experiments. However, seasonal differences in the production and processing of DOM were observed, reflecting differences in the nutrient regimes and initial DOM characteristics in each experiment, but also likely influenced by changes in the successional status of the pelagic community. Acknowledging that characteristics of the DOM derived from phytoplankton growth can vary broadly, heterotrophic processing and successional status of the community are synergistically important factors for shaping those characteristics, and thus affecting the seasonal signature of the semi-labile autochthonous DOM pool.
  • Elovaara, Samu; Eronen-Rasimus, Eeva; Asmala, Eero; Tamelander, Tobias; Kaartokallio, Hermanni (2021)
    Microbial consumption of phytoplankton-derived organic carbon in the pelagic food web is an important component of the global C cycle. We studied C cycling in two phytoplankton-bacteria systems (non-axenic cultures of a dinoflagellate Apocalathium malmogiense and a cryptophyte Rhodomonas marina) in two complementary experiments. In the first experiment we grew phytoplankton and bacteria in nutrient-replete conditions and followed C processing at early exponential growth phase and twice later when the community had grown denser. Cell-specific primary production and total community respiration were up to 4 and 7 times higher, respectively, in the A. malmogiense treatments. Based on the optical signals, accumulating dissolved organic C (DOC) was degraded more in the R. marina treatments, and the rate of bacterial production to primary production was higher. Thus, the flow of C from phytoplankton to bacteria was relatively higher in R. marina treatments than in A. malmogiense treatments, which was further supported by faster C-14 transfer from phytoplankton to bacterial biomass. In the second experiment we investigated consumption of the phytoplankton-derived DOC by bacteria. DOC consumption and transformation, bacterial production, and bacterial respiration were all higher in R. marina treatments. In both experiments A. malmogiense supported a bacterial community predominated by bacteria specialized in the utilization of less labile DOC (class Bacteroidia), whereas R. marina supported a community predominated by copiotrophic Alphaand Gammaproteobacteria. Our findings suggest that large dinoflagellates cycle relatively more C between phytoplankton biomass and the inorganic C pool, whereas small cryptophytes direct relatively more C to the microbial loop.
  • Pihan, Philippe; Lisbona, Fernanda; Borgonovo, Janina; Edwards-Jorquera, Sandra; Nunes-Hasler, Paula; Castillo, Karen; Kepp, Oliver; Urra, Hery; Saarnio, Suvi; Vihinen, Helena; Carreras-Sureda, Amado; Forveille, Sabrina; Sauvat, Allan; De Giorgis, Daniela; Pupo, Amaury; Rodriguez, Diego A.; Quarato, Giovanni; Sagredo, Alfredo; Lourido, Fernanda; Letai, Anthony; Latorre, Ramon; Kroemer, Guido; Demaurex, Nicolas; Jokitalo, Eija; Concha, Miguel L.; Glavic, Alvaro; Green, Douglas R.; Hetz, Claudio (2021)
    Programmed cell death is regulated by the balance between activating and inhibitory signals. Here, we have identified RECS1 (responsive to centrifugal force and shear stress 1) [also known as TMBIM1 (transmembrane BAX inhibitor motif containing 1)] as a proapoptotic member of the TMBIM family. In contrast to other proteins of the TMBIM family, RECS1 expression induces cell death through the canonical mitochondrial apoptosis pathway. Unbiased screening indicated that RECS1 sensitizes cells to lysosomal perturbations. RECS1 localizes to lysosomes, where it regulates their acidification and calcium content, triggering lysosomal membrane permeabilization. Structural modeling and electrophysiological studies indicated that RECS1 is a pH-regulated calcium channel, an activity that is essential to trigger cell death. RECS1 also sensitizes whole animals to stress in vivo in Drosophila melanogaster and zebrafish models. Our results unveil an unanticipated function for RECS1 as a proapoptotic component of the TMBIM family that ignites cell death programs at lysosomes.
  • Wang, Linping; Saarela, Jani; Poque, Sylvain; Valkonen, Jari P. T. (2020)
    The class 1 ribonuclease III (RNase III) encoded by Sweet potato chlorotic stunt virus (CSR3) suppresses RNA silencing in plant cells and thereby counters the host antiviral response by cleaving host small interfering RNAs, which are indispensable components of the plant RNA interference (RNAi) pathway. The synergy between sweet potato chlorotic stunt virus and sweet potato feathery mottle virus can reduce crop yields by 90%. Inhibitors of CSR3 might prove efficacious to counter this viral threat, yet no screen has been carried out to identify such inhibitors. Here, we report a novel high-throughput screening (HTS) assay based on fluorescence resonance energy transfer (FRET) for identifying inhibitors of CSR3. For monitoring CSR3 activity via HTS, we used a small interfering RNA substrate that was labelled with a FRET-compatible dye. The optimized HTS assay yielded 109 potential inhibitors of CSR3 out of 6,620 compounds tested from different small-molecule libraries. The three best inhibitor candidates were validated with a dose-response assay. In addition, a parallel screen of the selected candidates was carried out for a similar class 1 RNase III enzyme from Escherichia coli (EcR3), and this screen yielded a different set of inhibitors. Thus, our results show that the CSR3 and EcR3 enzymes were inhibited by distinct types of molecules, indicating that this HTS assay could be widely applied in drug discovery of class 1 RNase III enzymes.
  • Mõttus, Matti; Aragão, Luiz; Bäck, Jaana; Clemente, Rocío Hernandez; Maeda, Eduardo Eiji; Markiet, Vincent Robert Leon; Nichol, Caroline; Oliveira, Raimundo Cosme; Restrepo-Coupe, Natalia (2019)
    The spectral properties of plant leaves relate to the state of their photosynthetic apparatus and the surrounding environment. An example is the well known photosynthetic downregulation, active on the time scale from minutes to hours, caused by reversible changes in the xanthophyll cycle pigments. These changes affect leaf spectral absorption and are frequently quantified using the photochemical reflectance index (PRI). This index can be used to remotely monitor the photosynthetic status of vegetation, and allows for a global satellite-based measurement of photosynthesis. Such earth observation satellites in near-polar orbits usually cover the same geographical location at the same local solar time at regular intervals. To facilitate the interpretation of these instantaneous remote PRI measurements and upscale them temporally, we measured the daily course of leaf PRI in two evergreen biomes—a European boreal forest and an Amazon rainforest. The daily course of PRI was different for the two locations: At the Amazonian forest, the PRI of Manilkara elata leaves was correlated with the average photosynthetic photon flux density (PPFD) ( R2=0.59 ) of the 40 minutes preceding the leaf measurement. In the boreal location, the variations in Pinus sylvestris needle PRI were only weakly ( R2=0.27) correlated with mean PPFD of the preceding two hours; for Betula pendula, the correlation was insignificant regardless of the averaging period. The measured daily PRI curves were specific to species and/or environmental conditions. Hence, for a proper interpretation of satellite-measured instantaneous photosynthesis, the scaling of PRI measurements should be supported with information on its correlation with PPFD.
  • Valiev, R. R.; Nasibullin, R. T.; Cherepanov, V. N.; Baryshnikov, G.; Sundholm, D.; Ågren, H.; Minaev, B. F.; Kurten, T. (2020)
    A new method for calculating internal conversion rate constants (k_(IC)), including anharmonic effects and using the Lagrangian multiplier technique, is proposed. The deuteration effect on k_(IC) is investigated for naphthalene, anthracene, free-base porphyrin (H2P) and tetraphenylporphyrin (H2TPP). The results show that anharmonic effects are important when calculating k_(IC) for transitions between electronic states that are energetically separated (Delta E) by more than 20 000-25 000 cm(-1). Anharmonic effects are also important when Delta E <20 000-25 000 cm(-1) and when the accepting modes are X-H stretching vibrations with a frequency larger than 2000 cm(-1). The calculations show that there is mixing between the S-1 and S-2 states of naphthalene induced by non-adiabatic interactions. The non-adiabatic interaction matrix element between the S-1 and S-2 states is 250 cm(-1) and 50 cm(-1) for the normal and fully deuterated naphthalene structure and this difference significantly affects the estimated fluorescence quantum yield. Besides aromatic hydrocarbons H2P and H2TPP, the k_(IC) rate constant is also calculated for pyrometene (PM567) and tetraoxa[8]circulene (4B) with a detailed analysis of the effect of the vibrational anharmonicity.
  • Valiev, R. R.; Cherepanov, V. N.; Baryshnikov, G. V.; Sundholm, D. (2018)
    A method for calculating the rate constants for internal-conversion (k(IC)) and intersystem-crossing (k(ISC)) processes within the adiabatic and Franck-Condon (FC) approximations is proposed. The applicability of the method is demonstrated by calculation of k(IC) and k(ISC) for a set of organic and organometallic compounds with experimentally known spectroscopic properties. The studied molecules were pyrromethene-567 dye, psoralene, hetero[8]circulenes, free-base porphyrin, naphthalene, and larger polyacenes. We also studied fac-Alq(3) and fac-Ir(ppy)(3), which are important molecules in organic light emitting diodes (OLEDs). The excitation energies were calculated at the multi-configuration quasi-degenerate second-order perturbation theory (XMC-QDPT2) level, which is found to yield excitation energies in good agreement with experimental data. Spin-orbit coupling matrix elements, non-adiabatic coupling matrix elements, Huang-Rhys factors, and vibrational energies were calculated at the time-dependent density functional theory (TDDFT) and complete active space self-consistent field (CASSCF) levels. The computed fluorescence quantum yields for the pyrromethene-567 dye, psoralene, hetero[8]circulenes, fac-Alq(3) and fac-Ir(ppy)(3) agree well with experimental data, whereas for the free-base porphyrin, naphthalene, and the polyacenes, the obtained quantum yields significantly differ from the experimental values, because the FC and adiabatic approximations are not accurate for these molecules.
  • Silva, Luis; Calleja, Maria Ll.; Ivetic, Snjezana; Huete-Stauffer, Tamara; Roth, Florian; Carvalho, Susana; Moran, Xose Anxelu G. (2021)
    In coral reefs, dissolved organic matter (DOM) cycling is a critical process for sustaining ecosystem functioning. However, global and local stressors have caused persistent shifts from coral- to algae-dominated benthic communities. The influence of such phase shifts on DOM nature and its utilization by heterotrophic bacterioplankton remains poorly studied. Every second month for one year, we retrieved seawater samples enriched in DOM produced by coral- and algae-dominated benthic communities in a central Red Sea reef during a full annual cycle. Seawater incubations were conducted in the laboratory under in situ temperature and light conditions by inoculating enriched DOM samples with bacterial assemblages collected in the surrounding waters. Dissolved organic carbon (DOC) concentrations were higher in the warmer months (May-September) in both communities, resulting in higher specific growth rates and bacterial growth efficiencies (BGE). However, these high summer values were significantly enhanced in algal-DOM relative to coral-DOM, suggesting the potential for bacterioplankton biomass increase in reefs with algae replacing healthy coral cover under warmer conditions. The potential exacerbation of heterotrophic bacterial activity in the ongoing widespread regime shift from coral- to algae-dominated communities may have detrimental consequences for the overall health of tropical coral reefs. (C) 2020 The Authors. Published by Elsevier B.V.
  • Koivuniemi, Artturi; Fallarero, Adyary; Bunker, Alex (2019)
    The development of antimicrobial agents that target and selectively disrupt biofilms is a pressing issue since, so far, no antibiotics have been developed that achieve this effectively. Previous experimental work has found a promising set of antibacterial peptides: β2,2-amino acid derivatives, relatively small molecules with common structural elements composed of a polar head group and two non-polar hydrocarbon arms. In order to develop insight into possible mechanisms of action of these novel antibacterial agents, we have performed an in silico investigation of four leading β2,2-amino acid derivatives, interacting with models of both bacterial (target) and eukaryotic (host) membranes, using molecular dynamics simulation with a model with all-atom resolution. We found an unexpected result that could shed light on the mechanism of action of these antimicrobial agents: the molecules assume a conformation where one of the hydrophobic arms is directed downward into the membrane core while the other is directed upwards, out of the membrane and exposed above the position of the membrane headgroups; we dubbed this conformation the “can-can pose”. Intriguingly, the can-can pose was most closely linked to the choice of headgroup. Also, the compound previously found to be most effective against biofilms displayed the strongest extent of this behavior and, additionally, this behavior was more pronounced for this compound in the bacterial than in the eukaryotic membrane. We hypothesize that adopting the can-can pose could possibly disrupt the protective peptidoglycan macronet found on the exterior of the bacterial membrane.
  • Kerst, Thomas; Toivonen, Juha (2018)
    Luminescence in air induced by alpha particle emitters can be used to optically detect radioactive contamination from distances that surpass the range of the alpha radiation itself. Alpha particles excite nitrogen molecules in air and the relaxation creates a faint light emission. When the composition of the gases surrounding the alpha particle emitter is altered then the luminescence spectrum changes. In this work. we report the creation of an intense light emission in the wavelength regime below 300 nm originating from alpha particle excited nitric oxide (NO). The light yield has been investigated as a function of the NO concentration in an N-2 atmosphere. Unlike the emission from molecular nitrogen, NO emits at wavelengths shorter than 300 nm, where solar background and artificial lighting am negligible, thus enabling optical detection of alpha radiation even under bright lighting conditions. We show that the radioactively induced NO emission reaches its maximum intensity at a concentration of 50 ppm of NO diluted in N-2. At this concentration, the strongest emission line of NO is about 25 times more intense than the most intense line of N-2 radioluminescence. Lastly, we discuss potential applications and limitations of the technique. (C) 2018 Optical Society of America under the terms of the OSA Open Access Publishing Agreement
  • Rowe, Owen F.; Dinasquet, Julie; Paczkowska, Joanna; Figueroa, Daniela; Riemann, Lasse; Andersson, Agneta (2018)
    Dissolved organic matter (DOM) in marine waters is a complex mixture of compounds and elements that contribute substantially to the global carbon cycle. The large reservoir of dissolved organic carbon (DOC) represents a vital resource for heterotrophic bacteria. Bacteria can utilise, produce, recycle and transform components of the DOM pool, and the physicochemical characteristics of this pool can directly influence bacterial activity; with consequences for nutrient cycling and primary productivity. In the present study we explored bacterial transformation of naturally occurring DOM across an extensive brackish water gradient in the Baltic Sea. Highest DOC utilisation (indicated by decreased DOC concentration) was recorded in the more saline southerly region where waters are characterised by more autochthonous DOM. These sites expressed the lowest bacterial growth efficiency (BGE), whereas in northerly regions, characterised by higher terrestrial and allochthonous DOM, the DOC utilisation was low and BGE was highest. Bacterial processing of the DOM pool in the south resulted in larger molecular weight compounds and compounds associated with secondary terrestrial humic matter being degraded, and a processed DOM pool that was more aromatic in nature and contributed more strongly to water colour; while the opposite was true in the north. Nutrient concentration and stoichiometry and DOM characteristics affected bacterial activity, including metabolic status (BGE), which influenced DOM transformations. Our study highlights dramatic differences in DOM characteristics and microbial carbon cycling in sub-basins of the Baltic Sea. These findings are critical for our understanding of carbon and nutrient biogeochemistry, particularly in light of climate change scenarios.
  • Takamiya, Masanari; Xu, Feng; Suhonen, Heikki; Gourain, Victor; Yang, Lixin; Nga Yu Ho, [Tuntematon]; Helfen, Lukas; Schroeck, Anne; Etard, Christelle; Grabher, Clemens; Rastegar, Sepand; Schlunck, Guenther; Reinhard, Thomas; Baumbach, Tilo; Straehle, Uwe (2016)
    Altered levels of trace elements are associated with increased oxidative stress that is eventually responsible for pathologic conditions. Oxidative stress has been proposed to be involved in eye diseases, including cataract formation. We visualized the distribution of metals and other trace elements in the eye of zebrafish embryos by micro X-ray fluorescence (mu-XRF) imaging. Many elements showed highest accumulation in the retinal pigment epithelium (RPE) of the zebrafish embryo. Knockdown of the zebrafish brown locus homologues tyrp1a/b eliminated accumulation of these elements in the RPE, indicating that they are bound by mature melanosomes. Furthermore, albino (slc45a2) mutants, which completely lack melanosomes, developed abnormal lens reflections similar to the congenital cataract caused by mutation of the myosin chaperon Unc45b, and an in situ spin trapping assay revealed increased oxidative stress in the lens of albino mutants. Finally transplanting a wildtype lens into an albino mutant background resulted in cataract formation. These data suggest that melanosomes in pigment epithelial cells protect the lens from oxidative stress during embryonic development, likely by buffering trace elements.
  • Xu, Shan; Zaidan, Martha A.; Honkavaara, Eija; Hakala, Teemu; Viljanen, Niko; Porcar-Castell, Albert; Liu, Zhigang; Atherton, Jon (IEEE, 2020)
    IEEE International Symposium on Geoscience and Remote Sensing IGARSS
    Leaf angle distribution (LAD) is a key canopy structural parameter, playing an important role in light transfer. LAD can be estimated from fixed point of view photography, however this is time consuming and spatially limited. Recently, Terrestrial LiDAR Scanning (TLS) has been used to estimate LAD through 3D canopy space. The downside of TLS it is more costly than the cameras used in the photographic method. We propose a cost effective method to estimate LAD from drone based photogrammetry. We compare LAD estimates in different water treatment plots. Results show that LAD can be obtained from photogrammetric point clouds. Leaf angles were enhanced in stressed plots, presumably due to wilting. Further, the leaf azimuth distribution was not random but concentrated around 0 and 180 degrees. In summary, drone based photogrammetry can be used to estimate remote sensing parameters such as LAD paving the way for cost effective trait estimation.
  • Morales, Luis O.; Shapiguzov, Alexey; Safronov, Omid; Leppälä, Johanna; Vaahtera, Lauri; Yarmolinsky, Dmitry; Kollist, Hannes; Brosche, Mikael (2021)
    Tropospheric ozone (O-3) is a major air pollutant that decreases yield of important crops worldwide. Despite long-lasting research of its negative effects on plants, there are many gaps in our knowledge on how plants respond to O-3. In this study, we used natural variation in the model plant Arabidopsis (Arabidopsis thaliana) to characterize molecular and physiological mechanisms underlying O-3 sensitivity. A key parameter in models for O-3 damage is stomatal uptake. Here we show that the extent of O-3 damage in the sensitive Arabidopsis accession Shahdara (Sha) does not correspond with O-3 uptake, pointing toward stomata-independent mechanisms for the development of O-3 damage. We compared tolerant (Col-0) versus sensitive accessions (Sha, Cvi-0) in assays related to photosynthesis, cell death, antioxidants, and transcriptional regulation. Acute O-3 exposure increased cell death, development of lesions in the leaves, and decreased photosynthesis in sensitive accessions. In both Sha and Cvi-0, O-3-induced lesions were associated with decreased maximal chlorophyll fluorescence and low quantum yield of electron transfer from Photosystem II to plastoquinone. However, O-3-induced repression of photosynthesis in these two O-3-sensitive accessions developed in different ways. We demonstrate that O-3 sensitivity in Arabidopsis is influenced by genetic diversity given that Sha and Cvi-0 developed accession-specific transcriptional responses to O-3. Our findings advance the understanding of plant responses to O-3 and set a framework for future studies to characterize molecular and physiological mechanisms allowing plants to respond to high O-3 levels in the atmosphere as a result of high air pollution and climate change.
  • Göbüz, Göker; Liu, Chang; Jiang, Zhong-qing; Pulkkinen, Marjo; Piironen, Vieno; Sontag-Strohm, Tuula; Heinonen, Marina (2018)
    The course of protein-lipid co-oxidation was investigated in oil-in-water emulsions stabilized with proteins extracted from microwave-treated (MWT) and conventional thermal-treated (CTT) faba beans stored at 37 degrees C for 7days. Emulsions prepared with proteins from untreated (UT) faba beans and soy protein isolate (SP) were monitored for comparison. Lipid oxidation was detected through formation of primary and secondary oxidation products while protein oxidation was examined via tryptophan fluorescence degradation in interface and aqueous phase. Oxidation of proteins was more emphasized in the interfacial layers of MWT, CTT, and SP emulsions than in UT emulsions due to the prominence of radical chain-driven co-oxidation mechanism while lipoxygenase (LOX) activity in UT and MWT emulsions resulted in high amounts of hydroperoxides and abundance in lipid oxidation volatiles. Conventional thermal treatment provided better oxidative stability than microwave treatment reflected in lower levels of hydroperoxides and relative lack of diversity in lipid volatiles. Among detected volatiles, formation of ketones was more distinguished in MWT, CTT, and SP emulsions while UT emulsions contained a more diverse range of alkenals and alkanals. Ketones are known to form mainly through radical recombination reactions which combined with the results of protein oxidation supports that radical transfer reactions between proteins and lipids were the driving force behind oxidation in MWT, CTT, and SP emulsions. Treatments of faba beans resulted in increased oxidative stability of emulsified lipids and lower degradation of aqueous phase proteins.
  • Januszewski, Andrzej S.; Chen, David; Scott, Russell S.; O'Connell, Rachel L.; Aryal, Nanda R.; Sullivan, David R.; Watts, Gerald F.; Taskinen, Marja-Riitta; Barter, Philip J.; Best, James D.; Simes, R. John; Keech, Anthony C.; Jenkins, Alicia J. (2021)
    People with diabetes are at risk of chronic complications and novel biomarkers, such as Advanced glycation end-products (AGEs) may help stratify this risk. We assessed whether plasma low-molecular weight AGEs, also known as LMW-fluorophores (LMW-F), are associated with risk factors, predict complications, and are altered by fenofibrate in adults with type 2 diabetes. Plasma LMW-F were quantified at baseline, after six weeks fenofibrate, and one year post-randomisation to fenofibrate or placebo. LMW-F associations with existing and new composite vascular complications were determined, and effects of fenofibrate assessed. LMW-F correlated positively with age, glycated haemoglobin (HbA1c), pulse pressure, kidney dysfunction and inflammation; and negatively with urate, body mass index, oxidative stress and leptin, albeit weakly (r = 0.04-0.16, all p < 0.01). Independent determinants of LMW-F included smoking, diastolic blood pressure, prior cardiovascular disease or microvascular complications, Caucasian ethnicity, kidney function, HbA1c and diabetes duration (all p