Molecular patterns behind immunological and metabolic alterations in lysinuric protein intolerance

Lysinuric protein intolerance (LPI) is a recessively inherited disorder characterised by reduced plasma and increased urinary levels of cationic amino acids (CAAs), protein malnutrition, growth failure and hyperlipidemia. Some patients develop severe immunological, renal and pulmonary complications. All Finnish patients share the same LPIFin mutation in the SLC7A7 gene that encodes CAA transporter y+LAT1. The aim of this study was to examine molecular factors contributing to the various symptoms, systemic metabolic and lipid profiles, and innate immune responses in LPI. The transcriptomes, metabolomes and lipidomes were analysed in whole-blood cells and plasma using RNA microarrays and gas or liquid chromatography-mass spectrometry techniques, respectively. Toll-like receptor (TLR) signalling in monocyte-derived macrophages exposed to pathogens was scrutinised using qRT-PCR and the Luminex technology. Altered levels of transcripts participating in amino acid transport, immune responses, apoptosis and pathways of hepatic and renal metabolism were identified in the LPI whole-blood cells. The patients had increased non-essential amino acid, triacylglycerol and fatty acid levels, and decreased plasma levels of phosphatidylcholines and practically all essential amino acids. In addition, elevated plasma levels of eight metabolites, long-chain triacylglycerols, two chemoattractant chemokines and nitric oxide correlated with the reduced glomerular function in the patients with kidney disease. Accordingly, it can be hypothesised that the patients have increased autophagy, inflammation, oxidative stress and apoptosis, leading to hepatic steatosis, uremic toxicity and altered intestinal microbe metabolism. Furthermore, the LPI macrophages showed disruption in the TLR2/1, TLR4 and TLR9 pathways, suggesting innate immune dysfunctions with an excessive response to bacterial infections but a deficient viral DNA response.Lysinuurinen proteiini-intoleranssi (LPI) on peittyvästi periytyvä sairaus, jossa kationisten aminohappojen pitoisuudet ovat plasmassa matalat ja virtsassa korkeat ja potilailla esiintyy proteiinialiravitsemusta, kasvuhäiriöitä ja hyperlipidemiaa. Joillekin potilaille kehittyy lisäksi immunologisia sekä munuais- ja keuhkotoimintojen komplikaatioita. Kaikilla suomalaispotilailla on sama LPIFin-mutaatio SLC7A7-geenissä, joka koodaa kationisten aminohappojen kuljetinta y+LAT1:tä. Tämän tutkimuksen tarkoituksena oli selvittää molekulaaristen tekijöiden vaikutusta taudin moninaisiin oireisiin, systeemisiä metabolia- ja lipiditason muutoksia sekä synnynnäisen immuniteetin vasteita LPI-potilailla. Kokoveren soluista ja plasmasta analysoitiin RNA-mikrosiruja ja kaasu- tai nestekromatografia-massaspektrometriatekniikoita käyttämällä transkriptomit, metabolomit ja lipidomit. Tollinkaltaisten reseptorien (TLR) signalointia tarkasteltiin monosyyteistä erilaistetuissa patogeeneille altistuneissa makrofageissa käyttämällä qRT-PCR:ää ja Luminex-teknologiaa. Kokoveren soluista löydettiin aminohappokuljetukseen, immuunivasteisiin, apoptoosiin sekä maksa- ja munuaismetaboliareitteihin liittyviä transkripteja, joiden tasot olivat muuttuneet potilailla. Potilaiden ei-välttämättömien aminohappojen, triasyyliglyserolien ja rasvahappojen plasmapitoisuudet olivat kohonneet, kun taas fosfatidyylikoliinien ja lähes kaikkien välttämättömien aminohappojen pitoisuudet olivat alentuneet. Kahdeksan metaboliitin, pitkäketjuisten triasyyliglyserolien, kahden kemoatraktantin kemokiinin ja typpioksidin kohonneet plasmapitoisuudet korreloivat heikentyneen glomerulustoiminnan kanssa munuaistautia sairastavilla potilailla. Tulosten perusteella näyttää siltä, että LPI-potilailla on lisääntynyt autofagia, tulehdustila, oksidatiivinen stressi ja apoptoosi, jotka voivat johtaa maksan steatoosiin, toksisten aineiden kerääntymiseen veressä ja muuttuneeseen suolistomikrobien metaboliaan. Lisäksi LPI-makrofagien TLR2/1-, TLR4- ja TLR9-signaalivälitysreiteissä havaittiin muutoksia, jotka saattavat aiheuttaa synnynnäisen immuniteetin toimintahäiriöitä ja liiallisen vasteen bakteeri-infektioille mutta heikentyneen vasteen virus-DNA:lle.Siirretty Doriast

Developmental expression patterns of six6: a gene linked with spawning ecotypes in Atlantic salmon

The Atlantic salmon has been studied extensively, particularly as a model for understanding the genetic and environmental contributions to the evolution and development of life history traits. Expression pattern analysis in situ, however, is mostly lacking in salmon. We examine the embryonic developmental expression of six6, a candidate gene previously identified to be associated with spawning ecotypes and age at sexual maturity, in Atlantic salmon. Six6 is a member of the sine oculis homeobox family of transcription factors and is known to regulate eye and brain development in other vertebrates. We assay the expression of this gene in embryonic Atlantic salmon Salmo salar by whole-mount in situ hybridization. In line with earlier studies in other vertebrate species, we find conserved expression in the developing brain and sensory organs, including optic and olfactory primordia. However, we also find previously unreported domains of expression that suggest additional roles in axial and appendicular development, cardiovascular, intestinal, and sensory organogenesis. Each of these systems are important in the sensory ecology of Atlantic salmon, suggesting it is plausible that six6 may have pleiotropic roles in this complex phenotype.Peer reviewe

Sex-specific lipid profiles in the muscle of Atlantic salmon juveniles

Energy allocation in juvenile fish can have important implications for future life-history progression. Inherited and environmental factors determine when and where individuals allocate energy, and timely and sufficient energy reserves are crucial for reaching key life stages involved in the timing of maturation and sea migration. In Atlantic salmon, lipid reserves are predominantly found in the viscera and myosepta in the muscle and have been shown to play a key role in determining the timing of maturity. This life-history trait is tightly linked to fitness in many species and can be different between males and females, however, the details of relative energy allocation in juveniles of different sexes is not well understood. Therefore, the aim of this study was to investigate the effects of sex, genetics and environment during juvenile development of salmon on the amount and composition of their lipid reserves. To do so, juvenile salmon were fed one of two different lipid food contents during their first summer and autumn under common-garden conditions. Muscle lipid composition and concentrations were determined by thin layer chromatography. The muscle lipid class concentrations covaried negatively with body length and males showed higher concentrations than females for phosphatidylcholine, cholesterol, sphingomyelin, and triacylglycerol. This sex-specific difference in major lipid classes presents a new scope for understanding the regulation of lipids during juvenile development and gives direction for understanding how lipids may interact and influence major life-history traits in Atlantic salmon.Peer reviewe

Transcription Profiles of Age-at-Maturity-Associated Genes Suggest Cell Fate Commitment Regulation as a Key Factor in the Atlantic Salmon Maturation Process

Despite recent taxonomic diversification in studies linking genotype with phenotype, follow-up studies aimed at understanding the molecular processes of such genotype-phenotype associations remain rare. The age at which an individual reaches sexual maturity is an important fitness trait in many wild species. However, the molecular mechanisms regulating maturation timing processes remain obscure. A recent genome-wide association study in Atlantic salmon (Salmo salar) identified large-effect age-at-maturity-associated chromosomal regions including genes vgll3, akap11 and six6, which have roles in adipogenesis, spermatogenesis and the hypothalamic-pituitary-gonadal (HPG) axis, respectively. Here, we determine expression patterns of these genes during salmon development and their potential molecular partners and pathways. Using Nanostring transcription profiling technology, we show development- and tissue-specific mRNA expression patterns for vgll3, akap11 and six6. Correlated expression levels of vgll3 and akap11, which have adjacent chromosomal location, suggests they may have shared regulation. Further, vgll3 correlating with arhgap6 and yap1, and akap11 with lats1 and yap1 suggests that Vgll3 and Akap11 take part in actin cytoskeleton regulation. Tissue-specific expression results indicate that vgll3 and akap11 paralogs have sex-dependent expression patterns in gonads. Moreover, six6 correlating with slc38a6 and rtn1, and Hippo signaling genes suggests that Six6 could have a broader role in the HPG neuroendrocrine and cell fate commitment regulation, respectively. We conclude that Vgll3, Akap11 and Six6 may influence Atlantic salmon maturation timing via affecting adipogenesis and gametogenesis by regulating cell fate commitment and the HPG axis. These results may help to unravel general molecular mechanisms behind maturation.Peer reviewe

Maturation in Atlantic salmon (Salmo salar, Salmonidae) : a synthesis of ecological, genetic, and molecular processes

Over the past decades, Atlantic salmon (Salmo salar, Salmonidae) has emerged as a model system for sexual maturation research, owing to the high diversity of life history strategies, knowledge of trait genetic architecture, and their high economic value. The aim of this synthesis is to summarize the current state of knowledge concerning maturation in Atlantic salmon, outline knowledge gaps, and provide a roadmap for future work. We summarize the current state of knowledge: 1) maturation in Atlantic salmon takes place over the entire life cycle, starting as early as embryo development, 2) variation in the timing of maturation promotes diversity in life history strategies, 3) ecological and genetic factors influence maturation, 4) maturation processes are sex-specific and may have fitness consequences for each sex, 5) genomic studies have identified large-effect loci that influence maturation, 6) the brain-pituitary-gonadal axis regulates molecular and physiological processes of maturation, 7) maturation is a key component of fisheries, aquaculture, conservation, and management, and 8) climate change, fishing pressure, and other anthropogenic stressors likely have major effects on salmon maturation. In the future, maturation research should focus on a broader diversity of life history stages, including early embryonic development, the marine phase and return migration. We recommend studies combining ecological and genetic approaches will help disentangle the relative contributions of effects in different life history stages to maturation. Functional validation of large-effect loci should reveal how these genes influence maturation. Finally, continued research in maturation will improve our predictions concerning how salmon may adapt to fisheries, climate change, and other future challenges.Peer reviewe

Inhaled Sargramostim Induces Resolution of Pulmonary Alveolar Proteinosis in Lysinuric Protein Intolerance

Pulmonary alveolar proteinosis (PAP) is a potentially fatal complication of lysinuric protein intolerance (LPI), an inherited disorder of cationic amino acid transport. The patients often present with mild respiratory symptoms, which may rapidly progress to acute respiratory failure responding poorly to conventional treatment with steroids and bronchoalveolar lavations (BALs). The pathogenesis of PAP in LPI is still largely unclear. In previous studies, we have shown disturbances in the function and activity of alveolar macrophages of these patients, suggesting that increasing the activity and the number of macrophages by recombinant human GM-CSF (rhuGM-CSF) might be beneficial in this patient group.Two LPI patients with complicated PAP were treated with experimental inhaled rhuGM-CSF (sargramostim) after poor response to maximal conventional therapy. BAL fluid and cell samples from one patient were studied with light microscopy and transmission electron microscopy.Excellent response to therapy was observed in patient 1 with no compliance problems or side effects. Macrophages with myelin figure-like structures were seen in her BAL sample. Slight improvement of the pulmonary function was evident also in patient 2, but the role of sargramostim could not be properly evaluated due to the complicated clinical situation.In conclusion, inhaled rhuGM-CSF might be of benefit in patients with LPI-associated PAP.</p

Examining retail purchases of cigarettes and nicotine replacement therapy in Finland

IntroductionFinland's success in achieving the goal of its tobacco endgame largely depends on rectifying deficiencies in the delivery of smoking cessation services. One such weakness, which has not been documented with empirical data, is misuse of nicotine replacement therapy (NRT). This study's objective was to examine purchase patterns of NRT for estimating improper use of the medication. The study was based on the assumption that duration of a purchase episode is indicative of either proper use or misuse of NRT.MethodsThe participants (n=728), who purchased at least one NRT product in 2016 (mostly gum/lozenge), were selected through enrollment in a large customer loyalty program in Finland (LoCard). Participants were categorized into one of five groups according to their longest purchase episode of NRT, defined by purchases made in consecutive, 4-week intervals.ResultsMost participants, who did not adhere to NRT guidelines, either purchased the medication for too short (≤4 weeks, 63.5%) or too long (&gt;24 weeks, 13.2%) of a purchase episode. Median purchases of NRT in the first month of use were one and four in the former and latter, respectively. In contrast to other groups, persistent users (&gt;24 weeks) did not curtail purchases of NRT across several 4-week intervals, suggesting potential for dependence on NRT.ConclusionsThe observation that most purchase episodes were terminated prematurely is consistent with surveys reporting widespread NRT misuse. Given uncertainty of greater regulation of NRT sales through legislation, it would be prudent for Finnish retailers to promote proper use of the therapy

Gene co-expression patterns in Atlantic salmon adipose tissue provide a molecular link among seasonal changes, energy balance and age at maturity.

Sexual maturation in many fishes requires a major physiological change that involves a rapid transition between energy storage and usage. In Atlantic salmon, this transition for the initiation of maturation is tightly controlled by seasonality and requires a high-energy status. Lipid metabolism is at the heart of this transition since lipids are the main energy storing molecules. The balance between lipogenesis (lipid accumulation) and lipolysis (lipid use) determines energy status transitions. A genomic region containing a transcription co-factor of the Hippo pathway, vgll3, is the main determinant of maturation timing in Atlantic salmon. Interestingly, vgll3 acts as an inhibitor of adipogenesis in mice and its genotypes are potentially associated with seasonal heterochrony in lipid storage and usage in juvenile Atlantic salmon. Here, we explored changes in expression of more than 300 genes directly involved in the processes of adipogenesis, lipogenesis and lipolysis, as well as the Hippo pathway in the adipose tissue of immature and mature Atlantic salmon with distinct vgll3 genotypes. We found molecular evidence consistent with a scenario in which immature males with different vgll3 genotypes exhibit contrasting seasonal dynamics in their lipid profiles. We also identified components of the Hippo signalling pathway as potential major drivers of vgll3 genotype-specific differences in adipose tissue gene expression. This study demonstrates the importance of adipose gene expression patterns for directly linking environmental changes with energy balance and age at maturity through genetic factors bridging lipid metabolism, seasonality and sexual maturation