The Role of Oxysterol Binding Proteins in Macrophages

Biological membranes consist of several types of lipids that have important structural and functional roles. Moreover, these lipids can act as signaling molecules or affect the function of cellular proteins. Regulation of lipid homeostasis is critical for the maintenance of cellular physiology, and disturbances in lipid metabolism aspects are key components behind many diseases, including atherosclerosis. Macrophage cells play central roles in the development of atherosclerosis. The excessive uptake of modified lipoprotein particles in the vessel subendothelial space results in a proinflammatory response and the formation of macrophage foam cells. Oxysterols, oxidized cholesterol derivatives that primarily are taken up along with the modified lipoproteins, act at the crossroads of lipid metabolism and inflammation, and are important signaling lipids involved in atherogenesis. In this thesis the functions of three members of the Oxysterol Binding Protein (ORP) Family were studied in the macrophage cell type. Their role was studied by knocking down the proteins individually using lentiviruses. Moreover, the general function of these proteins was studied using other cell models. In the first part, the targeting of late endosomal protein ORP1L was investigated. ORP1L was shown to affect the motility and function of late endosomes in a sterol-dependent manner by mediating interactions between late endosomes and endoplasmic reticulum. Similar contacts between cellular compartments are also demonstrated for ORP3 and ORP8. In the second part the role of ORP8 in the macrophage was investigated by transcriptomic, biochemical and functional assays. ORP8 knockdown was shown to result in activation of several nuclear pathways and to affect cellular migration. In the third part, the lipid composition of macrophages was studied in conditions relevant for atherogenesis. The analysis revealed complex and profound changes in the cellular lipidome in the absence of ORPs, with implications for the inflammatory response, suggesting that ORP proteins modulate inflammatory processes. This work provides information on the cellular functions of macrophage-enriched ORP members, and reveals novel aspects in the function of the proteins. These data suggest that the modulation of ORP levels and/or function represents an attractive target for prevention or treatment of atherosclerosis.Biologiset kalvot koostuvat erilaisista lipideistä eli rasva-aineista, joilla on tärkeitä rakenteellisia ja toiminnallisia tehtäviä. Lipiditasapainon säätely on erittäin tärkeää solujen fysiologian kannalta, ja sen häiriöt ovat keskeisessä asemassa monien tautien, kuten valtimonkovettumataudin, taustalla. ---- Makrofaagit eli syöjäsolut ovat keskeisiä soluja valtimonkovettumataudin kehittymisen kannalta, sillä ne poistavat kolesterolia valtimoiden seinämistä. Liiallisena tämä prosessi voi johtaa tulehdusreaktioon ja makrofaagien muuttumiseen vaahtosoluiksi. Kolesterolin mukana makrofaagit ottavat sisäänsä oksisteroleja, hapettuneita kolesterolijohdannaisia. Oksisterolit ovat tehokkaita signalointilipidejä, jotka säätelevät solun tulehdusreaktioita sekä rasva-aineenvaihduntaa, ja niiden määrän on havaittu kasvavan valtimonkovettumataudin edetessä. Tässä väitöskirjatyössä on tutkittu kolmen oksisteroleja sitovan proteiiniperheen (ORP) jäsenen toimintaa. ORP1L, ORP3 ja ORP8 proteiinit ilmentyvät makrofaagi-solutyypissä erityisen runsaasti. Proteiinien toimintaa tutkittiin makrofaagi-solumallissa hiljentämällä proteiineja yksi kerrallaan käyttäen hyväksi lentiviruksia. Lisäksi proteiineja tutkittiin yleisemmin käyttäen hyväksi muita solumalleja ja tekniikoita. Tutkimuksissa havaittiin, että ORP1L proteiini säätelee myöhäisten endosomien liikettä ja toimintaa solun kolesterolistatuksesta riippuvaisesti muodostamalla vuorovaikutuksen soluelinten välille. Myös ORP3- ja ORP8-proteiinien havaittiin muodostavat vastaavia vuorovaikutuksia solun eri kalvostojen välillä. ORP8:n havaittiin säätelevän eri tumankuljetusreittejä, ja vaikuttavan makrofaagien liikkuvuuteen. Kolmannessa osatyössä tutkittiin ORP-proteiinien puuttumisen vaikutusta makrofaagien lipidikoostumukseen olosuhteissa, jotka ovat olennaisia valtimonkovettumataudin kehittymisen kannalta. Proteiinien puutos aiheutti huomattavia muutoksia, mm. tulehduksellisissa lipideissä. Tulokset viittaavat siihen, että ORP-proteiineilla säätelevät makrofaagien tulehdusreaktioita. Tutkimus on antanut tärkeää uutta tietoa ORP-proteiinien toiminnasta sekä niiden vaikutuksista solujen fysiologian kannalta. Tutkimuksen perusteella voidaan sanoa, että ORP-proteiinien tasojen ja/tai toiminnan säätelystä saattaa olla hyötyä valtimonkovettumataudin ennaltaehkäisyssä tai hoidossa

Comparison of various strategies to define the optimal target population for liver fibrosis screening : A population-based cohort study

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OSBP-Related Protein 8 (ORP8) Regulates Plasma and Liver Tissue Lipid Levels and Interacts with the Nucleoporin Nup62

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Gender, Contraceptives and Individual Metabolic Predisposition Shape a Healthy Plasma Lipidome

Lipidomics of human blood plasma is an emerging biomarker discovery approach that compares lipid profiles under pathological and physiologically normal conditions, but how a healthy lipidome varies within the population is poorly understood. By quantifying 281 molecular species from 27 major lipid classes in the plasma of 71 healthy young Caucasians whose 35 clinical blood test and anthropometric indices matched the medical norm, we provided a comprehensive, expandable and clinically relevant resource of reference molar concentrations of individual lipids. We established that gender is a major lipidomic factor, whose impact is strongly enhanced by hormonal contraceptives and mediated by sex hormone-binding globulin. In lipidomics epidemiological studies should avoid mixed-gender cohorts and females taking hormonal contraceptives should be considered as a separate sub-cohort. Within a gender-restricted cohort lipidomics revealed a compositional signature that indicates the predisposition towards an early development of metabolic syndrome in ca. 25% of healthy male individuals suggesting a healthy plasma lipidome as resource for early biomarker discovery

hiPSC-derived hepatocytes closely mimic the lipid profile of primary hepatocytes: A future personalised cell model for studying the lipid metabolism of the liver

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Roadmap for implementing environmental DNA (eDNA) and other molecular monitoring methods in Finland – Vision and action plan for 2022–2025

Roadmap for implementing environmental DNA (eDNA) and other molecular monitoring methods in Finland – Vision and Action Plan for 2022-2025 Technological development in molecular methodology has been extremely fast in the past two decades, and groundbreaking new approaches have been introduced. It is now possible to detect and quantify DNA or RNA of target species or even map the whole species community in environmental samples of water, sediment, soil, air or assemblages of whole organisms. Moreover, the costs of high-throughput sequencing and other advanced molecular methods have decreased and methodological pipelines from sampling to data analysis developed sufficiently to allow large-scale, routine application of the new methods in environmental monitoring. This presents a huge opportunity to improve the coverage, accuracy and cost-efficiency of monitoring, enabling a much more complete picture of biodiversity and the state of the environment and their trends. As the new European Biodiversity Strategy for 2030 and other international policies to halt biodiversity loss and the degradation of habitats are translated into concrete measures, the quality of the monitoring data will play a crucial role in determining their success or failure. In this roadmap commissioned by the Finnish Ministry of the Environment, we assess the state-of-the-art in molecular monitoring methods in Finland within the international context, identify challenges and development areas that remain to be addressed and propose an action plan for promoting the coordinated implementation of molecular methods in national monitoring programs. Apart from the most recent scientific literature, our analysis is based on survey results, direct enquiries and interviews. Participation of the national community of experts from different sectors was enabled and invited at several stages of the roadmap preparation. Internationally, molecular monitoring methods are being actively developed and are routinely implemented in monitoring across different taxa and ecosystems. In Finland, molecular monitoring methods have been tested and piloted by all major institutions responsible for environmental monitoring, and the methods are already applied routinely in the monitoring of individual game species such as the wolf and European and Canadian beaver. However, other areas such as the monitoring of biodiversity, threatened species, non-mammalian invasive species or emerging plant or animal pests remain less developed, and national efforts and expertise are scattered across different organizations. Funding and know-how are perceived as the most important factors limiting molecular monitoring method implementation. We estimate that extensive, routine implementation of a wide range of molecular monitoring methods is conceivable in Finland before 2030. As the primary development areas for reaching this goal, we identify (i) international coordination and standard development, (ii) networking across sectors, (iii) education, (iv) infrastructure, (v) reference sequence libraries and the mapping of whole genomes, and (vi) modelling and analysis tool development. For concrete actions in 2022–2025, we propose (1) a cross-governmental funding instrument, (2) a permanent working group responsible for national and international coordination, (3) a national network and (4) an online platform to enhance interaction and knowledge transfer, as well as (5) a national data management system with collectively agreed data and metadata formats and standards. ---------- Kansallinen tiekartta ympäristö-DNA:n ja muiden molekyylibiologisten seurantamenetelmien käyttöönotolle – visio ja toimenpidesuunnitelma vuosille 2022-2025 Molekyylibiologisten menetelmien teknologinen kehitys on ollut ennennäkemättömän nopeaa kahden viime vuosikymmenen aikana. Uudet menetelmät mahdollistavat kohdelajien DNA:n tai RNA:n havaitsemisen ja runsausmäärityksen tai koko eliöyhteisön kartoittamisen esimerkiksi vesi-, sedimentti-, maaperä- tai ilmanäytteistä tai kokonaisia yksilöitä sisältävistä kokoomanäytteistä. Massiivisen rinnakkaissekvensoinnin ja muiden menetelmien kustannukset ovat merkittävästi laskeneet ja menetelmäketjut näytteenotosta tulosten tulkintaan kehittyneet asteelle, joka mahdollistaa niiden laajamittaisen, rutiininomaisen käytön ympäristön seurannassa. Uusien menetelmien avulla voimme parantaa seurannan kattavuutta, tarkkuutta ja kustannustehokkuutta ja siten täydentää seurannan kautta muodostuvaa kuvaa luonnon monimuotoisuudesta ja sen muutoksista. Tälle tiedolle on suuri tarve – laadukas seuranta on keskeinen edellytys sille, että EU:n uuden biodiversiteettistrategian ja muiden luontokadon ja elinympäristöjen tilan huonontumisen pysäyttämiseen tähtäävien kansainvälisten sitoumusten toimeenpano onnistuu. Tässä ympäristöministeriön tilaamassa tiekartassa arvioimme molekyylibiologisten seurantamenetelmien nykytilaa Suomessa osana laajempaa kansainvälistä kenttää, tunnistamme huomiota vaativia haasteita ja kehityskohteita ja ehdotamme konkreettisia toimenpiteitä molekyylibiologisten seurantamenetelmien koordinoidun käyttöönoton edistämiseksi lähivuosien aikana. Selvityksemme perustuu uusimman tieteellisen kirjallisuuden lisäksi kyselytutkimukseen sekä suoriin tiedusteluihin ja haastatteluihin. Yhteiskunnan eri sektoreita edustava kansallinen asiantuntijayhteisö osallistui tiekartan valmisteluun työn eri vaiheissa. Molekyylibiologisia seurantamenetelmiä kehitetään parhaillaan aktiivisesti ympäri maailmaa eri eliöryhmille ja ekosysteemeille, ja yksittäisiä menetelmiä on useissa maissa otettu myös rutiininomaiseen käyttöön. Suomessa menetelmiä on kehitetty ja pilotoitu kaikissa keskeisissä ympäristön seurantaa koordinoivissa laitoksissa, ja yksittäisten riistaeläinten kuten suden ja kanadan- ja euroopanmajavan seurannassa ne ovat jo rutiinikäytössä. Biodiversiteetin, uhanalaisten lajien, vieraslajien (nisäkkäitä lukuun ottamatta) ja muiden haitallisten lajien kansallisessa seurannassa molekyylibiologisten menetelmien käyttö on kuitenkin vielä kokeiluasteella, ja kehittämishankkeiden ja asiantuntijuuden kenttä on hajanainen. Riittämätöntä rahoitusta ja osaamista pidetään alan asiantuntijoiden keskuudessa tärkeimpinä menetelmien käyttöönottoa rajoittavina tekijöinä. Arviomme mukaan laaja kirjo molekyylibiologisia seurantamenetelmiä olisi mahdollista ottaa laajamittaiseen rutiininomaiseen käyttöön vuoteen 2030 mennessä. Tärkeimmiksi kehityskohteiksi nousivat (i) kansainvälinen koordinaatio ja menetelmien standardointi, (ii) organisaatioiden ja sektoreiden välinen verkostoituminen, (iii) koulutus, (iv) infrastruktuuri, (v) referenssisekvenssikirjastot ja kokonaisten genomien kartoittaminen sekä (vi) malli- ja analyysityökalujen kehittäminen. Konkreettisiksi toimenpiteiksi vuosille 2022-2025 esitämme (1) poikkihallinnollista rahoitusohjelmaa molekyylibiologisten seurantamenetelmien käyttöönottoa edistäville tutkimus- ja kehityshankkeille, (2) pysyvää työryhmää kansallisen ja kansainvälisen koordinaation edistämiseksi, (3) olemassa olevan kansallisen asiantuntijaverkoston laajentamista, (4) internet-pohjaista alustaa vuorovaikutuksen ja tiedonjaon tehostamiseksi sekä (5) kansallista, yhdessä sovittuja data- ja metadatastandardeja noudattavaa molekyylibiologisten seuranta-aineistojen tiedonhallintajärjestelmää

Roadmap for implementing environmental DNA (eDNA) and other molecular monitoring methods in Finland–Vision and action plan for 2022–2025

Technological development in molecular methodology has been extremely fast in the past two decades, and groundbreaking new approaches have been introduced. It is now possible to detect and quantify DNA or RNA of target species or even map the whole species community in environmental samples of water, sediment, soil, air or assemblages of whole organisms. Moreover, the costs of high-throughput sequencing and other advanced molecular methods have decreased and methodological pipelines from sampling to data analysis developed sufficiently to allow large-scale, routine application of the new methods in environmental monitoring. This presents a huge opportunity to improve the coverage, accuracy and cost-efficiency of monitoring, enabling a much more complete picture of biodiversity and the state of the environment and their trends. As the new European Biodiversity Strategy for 2030 and other international policies to halt biodiversity loss and the degradation of habitats are translated into concrete measures, the quality of the monitoring data will play a crucial role in determining their success or failure. In this roadmap commissioned by the Finnish Ministry of the Environment, we assess the state-ofthe-art in molecular monitoring methods in Finland within the international context, identify challenges and development areas that remain to be addressed and propose an action plan for promoting the coordinated implementation of molecular methods in national monitoring programs. Apart from the most recent scientific literature, our analysis is based on survey results, direct enquiries and interviews. Participation of the national community of experts from different sectors was enabled and invited at several stages of the roadmap preparation. Internationally, molecular monitoring methods are being actively developed and are routinely implemented in monitoring across different taxa and ecosystems. In Finland, molecular monitoring methods have been tested and piloted by all major institutions responsible for environmental monitoring, and the methods are already applied routinely in the monitoring of individual game species such as the wolf and European and Canadian beaver. However, other areas such as the monitoring of biodiversity, threatened species, non-mammalian invasive species or emerging plant or animal pests remain less developed, and national efforts and expertise are scattered across different organizations. Funding and know-how are perceived as the most important factors limiting molecular monitoring method implementation. We estimate that extensive, routine implementation of a wide range of molecular monitoring methods is conceivable in Finland before 2030. As the primary development areas for reaching this goal, we identify (i) international coordination and standard development, (ii) networking across sectors, (iii) education, (iv) infrastructure, (v) reference sequence libraries and the mapping of whole genomes, and (vi) modelling and analysis tool development. For concrete actions in 2022–2025, we propose (1) a cross-governmental funding instrument, (2) a permanent working group responsible for national and international coordination, (3) a national network and (4) an online platform to enhance interaction and knowledge transfer, as well as (5) a national data management system with collectively agreed data and metadata formats and standards

Developing a spatially explicit modelling and evaluation framework for integrated carbon sequestration and biodiversity conservation: application in southern Finland

The challenges posed by climate change and biodiversity loss are deeply interconnected. Successful co-managing of these tangled drivers requires innovative methods that can prioritize and target management actions against multiple criteria, while also enabling cost-effective land use planning and impact scenario assessment. This paper synthesises the development and application of an integrated multidisciplinary modelling and evaluation framework for carbon and biodiversity in forest systems. By analysing and spatio-temporally modelling carbon processes and biodiversity elements, we determine an optimal solution for their co-management in the study landscape. We also describe how advanced Earth Observation measurements can be used to enhance mapping and monitoring of biodiversity and ecosystem processes. The scenarios used for the dynamic models were based on official Finnish policy goals for forest management and climate change mitigation. The development and testing of the system were executed in a large region in southern Finland (Kokemäenjoki basin, 27 024 km2) containing highly instrumented LTER (Long-Term Ecosystem Research) stations; these LTER data sources were complemented by fieldwork, remote sensing and national data bases. In the study area, estimated total net emissions were currently 4.2 TgCO2eq a-1, but modelling of forestry measures and anthropogenic emission reductions demonstrated that it would be possible to achieve the stated policy goal of carbon neutrality by low forest harvest intensity. We show how this policy-relevant information can be further utilised for optimal allocation of set-aside forest areas for nature conservation, which would significantly contribute to preserving both biodiversity and carbon values in the region. Biodiversity gain in the area could be increased without a loss of carbon-related benefits.The challenges posed by climate change and biodiversity loss are deeply interconnected. Successful co-managing of these tangled drivers requires innovative methods that can prioritize and target management actions against multiple criteria, while also enabling cost-effective land use planning and impact scenario assessment. This paper synthesises the development and application of an integrated multidisciplinary modelling and evaluation framework for carbon and biodiversity in forest systems. By analysing and spatio-temporally modelling carbon processes and biodiversity elements, we determine an optimal solution for their co-management in the study landscape. We also describe how advanced Earth Observation measurements can be used to enhance mapping and monitoring of biodiversity and ecosystem processes. The scenarios used for the dynamic models were based on official Finnish policy goals for forest management and climate change mitigation. The development and testing of the system were executed in a large region in southern Finland (Kokemäenjoki basin, 27,024 km2) containing highly instrumented LTER (Long-Term Ecosystem Research) stations; these LTER data sources were complemented by fieldwork, remote sensing and national data bases. In the study area, estimated total net emissions were currently 4.2 TgCO2eq a−1, but modelling of forestry measures and anthropogenic emission reductions demonstrated that it would be possible to achieve the stated policy goal of carbon neutrality by low forest harvest intensity. We show how this policy-relevant information can be further utilized for optimal allocation of set-aside forest areas for nature conservation, which would significantly contribute to preserving both biodiversity and carbon values in the region. Biodiversity gain in the area could be increased without a loss of carbon-related benefits.Peer reviewe

Plasma ceramides predict cardiovascular death in patients with stable coronary artery disease and acute coronary syndromes beyond LDL-cholesterol

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