Plant cell cultures as food-aspects of sustainability and safety.

KEY MESSAGE: Sustainability and safety aspects of plant cell cultures as food are presented. Applicability of dairy side streams as carbon source and use of natural growth enhancers in cultivation are shown. Biotechnologically produced cellular products are currently emerging to replace and add into the portfolio of agriculturally derived commodities. Plant cell cultures used for food could supplement current food production. However, still many aspects need to be resolved before this new food concept can enter the market. Issues related to sustainability and safety for human consumption are relevant for both consumers and regulators. In this study, two plant cell cultures, deriving from arctic bramble (Rubus arcticus) and birch (Betula pendula), were cultivated using lactose-rich dairy side streams as alternative carbon sources to replace sucrose. Biomasses were comparable to those of original plant cell culture media when up to 83% and 75% of the original sucrose was replaced by these side streams for arctic bramble and birch cell cultures, respectively. Furthermore, nutritional composition or sensory properties were not compromised. Synthetic plant growth regulators were replaced by natural components, such as coconut water and IAA for several subculture cycles. Finally, it was shown that only trace amounts of free growth regulators are present in the cells at the harvesting point and assessment by freshwater crustaceans assay indicated that toxicity of the cells was not exceeding that of traditionally consumed bilberry fruit.

Biotransformation of Cyclodextrine-Complexed Semisynthetic Betulin Derivatives by Plant Cells.

In this study, three semisynthetic betulonic acid-based compounds, 20(29)-dihydrolup-2-en[2,3-d]isoxazol-28-oic acid, 1-betulonoylpyrrolidine, and lupa-2,20(29)-dieno[2,3-b]pyrazin-28-oic acid, were studied in biotransformation experiments using Nicotiana tabacum and Catharanthus roseus cell suspension cultures. Biotransformation was performed using cyclodextrin to aid dissolving poorly water-soluble substrates. Several new derivatives were found, consisting of oxidized and glycosylated (pentose- and hexose-conjugated) products.

Interaction between dietary lipids and gut microbiota regulates hepatic cholesterol metabolism.

The gut microbiota influences many aspects of host metabolism. We have previously shown that the presence of a gut microbiota remodels lipid composition. Here we investigated how interaction between gut microbiota and dietary lipids regulates lipid composition in the liver and plasma, and gene expression in the liver. Germ-free and conventionally raised mice were fed a lard or fish oil diet for 11 weeks. We performed lipidomics analysis of the liver and serum and microarray analysis of the liver. As expected, most of the variation in the lipidomics dataset was induced by the diet, and abundance of most lipid classes differed between mice fed lard and fish oil. However, the gut microbiota also affected lipid composition. The gut microbiota increased hepatic levels of cholesterol and cholesteryl esters in mice fed lard, but not in mice fed fish oil. Serum levels of cholesterol and cholesteryl esters were not affected by the gut microbiota. Genes encoding enzymes involved in cholesterol biosynthesis were downregulated by the gut microbiota in mice fed lard and were expressed at a low level in mice fed fish oil independent of microbial status. In summary, we show that gut microbiota-induced regulation of hepatic cholesterol metabolism is dependent on dietary lipid composition.

A Healthy Nordic Diet Alters the Plasma Lipidomic Profile in Adults with Features of Metabolic Syndrome in a Multicenter Randomized Dietary Intervention.

BACKGROUND: A healthy Nordic diet is associated with improvements in cardiometabolic risk factors, but the effect on lipidomic profile is not known. OBJECTIVE: The aim was to investigate how a healthy Nordic diet affects the fasting plasma lipidomic profile in subjects with metabolic syndrome. METHODS: Men and women (n = 200) with features of metabolic syndrome [mean age: 55 y; body mass index (in kg/m2): 31.6] were randomly assigned to either a healthy Nordic (n = 104) or a control (n = 96) diet for 18 or 24 wk at 6 centers. Of the participants, 156 completed the study with plasma lipidomic measurements. The healthy Nordic diet consisted of whole grains, fruits, vegetables, berries, vegetable oils and margarines, fish, low-fat milk products, and low-fat meat. An average Nordic diet served as the control diet and included low-fiber cereal products, dairy fat-based spreads, regular-fat milk products, and a limited amount of fruits, vegetables, and berries. Lipidomic profiles were measured at baseline, week 12, and the end of the intervention (18 or 24 wk) by using ultraperformance liquid chromatography mass spectrometry. The effects of the diets on the lipid variables were analyzed with linear mixed-effects models. Data from centers with 18- or 24-wk duration were also analyzed separately. RESULTS: Changes in 21 plasma lipids differed significantly between the groups at week 12 (false discovery rate P < 0.05), including increases in plasmalogens and decreases in ceramides in the healthy Nordic diet group compared with the control group. At the end of the study, changes in lipidomic profiles did not differ between the groups. However, when the intervention lasted 24 wk, changes in 8 plasma lipids that had been identified at 12 wk, including plasmalogens, were sustained. There were no differences in changes in plasma lipids between groups with an intervention of 18 wk. By the dietary biomarker score, adherence to diet did not explain the difference in the results related to the duration of the study. CONCLUSIONS: A healthy Nordic diet transiently modified the plasma lipidomic profile, specifically by increasing the concentrations of antioxidative plasmalogens and decreasing insulin resistance-inducing ceramides. This trial was registered at clinicaltrials.gov as NCT00992641.

Association of lipidome remodeling in the adipocyte membrane with acquired obesity in humans.

Identification of early mechanisms that may lead from obesity towards complications such as metabolic syndrome is of great interest. Here we performed lipidomic analyses of adipose tissue in twin pairs discordant for obesity but still metabolically compensated. In parallel we studied more evolved states of obesity by investigating a separated set of individuals considered to be morbidly obese. Despite lower dietary polyunsaturated fatty acid intake, the obese twin individuals had increased proportions of palmitoleic and arachidonic acids in their adipose tissue, including increased levels of ethanolamine plasmalogens containing arachidonic acid. Information gathered from these experimental groups was used for molecular dynamics simulations of lipid bilayers combined with dependency network analysis of combined clinical, lipidomics, and gene expression data. The simulations suggested that the observed lipid remodeling maintains the biophysical properties of lipid membranes, at the price, however, of increasing their vulnerability to inflammation. Conversely, in morbidly obese subjects, the proportion of plasmalogens containing arachidonic acid in the adipose tissue was markedly decreased. We also show by in vitro Elovl6 knockdown that the lipid network regulating the observed remodeling may be amenable to genetic modulation. Together, our novel approach suggests a physiological mechanism by which adaptation of adipocyte membranes to adipose tissue expansion associates with positive energy balance, potentially leading to higher vulnerability to inflammation in acquired obesity. Further studies will be needed to determine the cause of this effect.

Quantitative profiling of bile acids in blood, adipose tissue, intestine, and gall bladder samples using ultra high performance liquid chromatography-tandem mass spectrometry.

An ultra high performance liquid chromatography tandem mass spectrometry method (UHPLC-MS/MS) was developed for the determination of 33 target and 28 unknown bile acids (BAs) in biological samples. Sixty-one BAs could be measured in 20 min using only a small amount of sample and with a simple sample preparation. The method proved to be very sensitive (limit of detection 5-350 pg/mL, lower limit of quantitation 0.1-2.6 ng/mL), linear (R(2) > 0.99) and reproducible (typically CV <15 % in biological matrixes). The method was used to analyze human adipose tissue, plasma, and serum (from same subjects) and mouse serum, gall bladder, small intestine, and colon samples (from same animals). Cholic acid, ursodeoxycholic acid, and chenodeoxycholic acid, deoxycholic acid, and their conjugates (mainly glycine, but also taurine conjugates) were the main metabolites in human samples, and cholic acid, glycine cholic acid, and several taurine conjugates in mouse samples. Using the method, 28 unknown BAs could also be detected. UHPLC-MS/MS spectra, accurate mass, and tissue distribution suggested that nine of the unknown bile acids were taurine conjugates, 13 were glycine conjugates, and six were intact BAs, respectively. To our knowledge, this was the first time BAs were detected in adipose tissue. Results showed that 17 targeted BAs were found at ng/g level in human adipose tissue. Our findings give a novel insight of the endogenous role of BAs in adipose tissue and their role as biomarkers (e.g., in metabolic diseases).

Prediction of non-alcoholic fatty-liver disease and liver fat content by serum molecular lipids.

AIMS/HYPOTHESIS: We examined whether analysis of lipids by ultra-performance liquid chromatography (UPLC) coupled to MS allows the development of a laboratory test for non-alcoholic fatty-liver disease (NAFLD), and how a lipid-profile biomarker compares with the prediction of NAFLD and liver-fat content based on routinely available clinical and laboratory data. METHODS: We analysed the concentrations of molecular lipids by UPLC-MS in blood samples of 679 well-characterised individuals in whom liver-fat content was measured using proton magnetic resonance spectroscopy ((1)H-MRS) or liver biopsy. The participants were divided into biomarker-discovery (n = 287) and validation (n = 392) groups to build and validate the diagnostic models, respectively. RESULTS: Individuals with NAFLD had increased triacylglycerols with low carbon number and double-bond content while lysophosphatidylcholines and ether phospholipids were diminished in those with NAFLD. A serum-lipid signature comprising three molecular lipids ('lipid triplet') was developed to estimate the percentage of liver fat. It had a sensitivity of 69.1% and specificity of 73.8% when applied for diagnosis of NAFLD in the validation series. The usefulness of the lipid triplet was demonstrated in a weight-loss intervention study. CONCLUSIONS/INTERPRETATION: The liver-fat-biomarker signature based on molecular lipids may provide a non-invasive tool to diagnose NAFLD, in addition to highlighting lipid molecular pathways involved in the disease.

Inducible Synthetic Growth Regulation Using the ClpXP Proteasome Enhances cis,cis-Muconic Acid and Glycolic Acid Yields in Saccharomyces cerevisiae.

Engineered microbial cells can produce sustainable chemistry, but the production competes for resources with growth. Inducible synthetic control over the resource use would enable fast accumulation of sufficient biomass and then divert the resources to production. We developed inducible synthetic resource-use control overSaccharomyces cerevisiae by expressing a bacterial ClpXP proteasome from an inducible promoter. By individually targeting growth-essential metabolic enzymes Aro1, Hom3, and Acc1 to the ClpXP proteasome, cell growth could be efficiently repressed during cultivation. The ClpXP proteasome was specific to the target proteins, and there was no reduction in the targets when ClpXP was not induced. The inducible growth repression improved product yields from glucose (cis,cis-muconic acid) and per biomass (cis,cis-muconic acid and glycolic acid). The inducible ClpXP proteasome tackles uncertainties in strain optimization by enabling model-guided repression of competing, growth-essential, and metabolic enzymes. Most importantly, it allows improving production without compromising biomass accumulation when uninduced; therefore, it is expected to mitigate strain stability and low productivity challenges.

Liquid Chromatography-Mass Spectrometry (LC-MS)-Based Analysis of Molecular Lipids in Algae Samples.

In this paper, an analytical method for the analysis of molecular lipids in algae samples is reported. The sample preparation is based on a modified Folch extraction, and the analysis is carried out with ultrahigh performance liquid chromatography combined with mass spectrometry (UPLC-MS). For further characterization of lipids, MS/MS analyses are carried out utilizing either a separate instrument (e.g., LTQ-Orbitrap mass spectrometer) or simultaneous fragmentation with the same instrument. Throughput of the method is over 100 samples/d. The repeatability is good, and the relative standard deviation of spiked samples is <15%.

UPLC-ELSD Analysis of Algal Lipid Classes and Derivatization of Bound and Free Fatty Acids and Sterols for GC-MS Methods.

Constituents of microalgae and sample preparation for UPLC-ELSD and GC-MS analyses are described. Bound fatty acids from acylglycerols, alkylacylglycerols, galactosyldiacylglycerols, glycerophospholipids, and sterol esters are derivatized by using transesterification with sodium methoxide to form fatty acid methyl esters. Compounds containing free hydroxyl groups, either present originally or formed during previous step, like free fatty acids, sterols, α-tocopherol, phytol, and nonesterified alkoxyglycerols, are trimethylsilylated. The compounds in algal lipid extract are subsequently derivatized by these two steps.

Examining Safety of Biocolourants from Fungal and Plant Sources-Examples from Cortinarius and Tapinella, Salix and Tanacetum spp. and Dyed Woollen Fabrics.

Biocolourants have been investigated as alternatives to synthetic dyes. However, natural origin per se is not a label of harmlessness and research is needed to obtain safe dyes. We studied the cytotoxicity of the extracts from fungal (Cortinarius semisanguineus, Tapinella atrotomentosa) and plant (Tanacetum vulgare, Salix phylicifolia) sources and the woollen fabrics dyed with the extracts. Cytotoxicity in vitro using hepa-1 mouse hepatoma cells for 24 h and 72 h exposure was observed as the highest tolerated dose. All biocolourants produced intensive colour on fabrics with fastness properties from moderate to good. The Salix and Cortinarius samples did not show any cytotoxic effects, whereas the Tanacetum and Tapinella samples had slightly higher test values but were not interpreted as being significantly toxic. Higher than zero values of the undyed fabrics showed the importance of examining their toxicity as well. It was found that the cytotoxicity of the samples dyed with the biocolourants did not differ significantly from the undyed wool fabric. The concentrations of dyes used in the assays were very low, imitating the dose of the user. In addition to colouring properties, natural dyes may have pharmaceutical and antibacterial properties which would enhance the interest in using them in products for added value.

Plant cells as food - A concept taking shape.

Plant cell cultures from cloudberry, lingonberry and stoneberry were studied in terms of their nutritional properties as food. Carbohydrate, lipid and protein composition, in vitro protein digestibility and sensory properties were investigated. Dietary fibre content varied between 21.2 and 36.7%, starch content between 0.3 and 1.3% and free sugar content between 17.6 and 33.6%. Glucose and fructose were the most abundant sugars. High protein contents between 13.7 and 18.9% were recorded and all samples had a balanced amino acid profile. In vitro protein digestion assay showed hydrolysis by digestive enzymes in fresh cells but only limited hydrolysis in freeze-dried samples. The lipid analysis indicated that the berry cells were rich sources of essential, polyunsaturated fatty acids. In sensory evaluation, all fresh berry cells showed fresh odour and flavour. Fresh cell cultures displayed a rather sandy, coarse mouthfeel, whereas freeze-dried cells melted quickly in the mouth. All in all the potential of plant cells as food was confirmed.

Genome wide analysis of protein production load in Trichoderma reesei.

BACKGROUND: The filamentous fungus Trichoderma reesei (teleomorph Hypocrea jecorina) is a widely used industrial host organism for protein production. In industrial cultivations, it can produce over 100 g/l of extracellular protein, mostly constituting of cellulases and hemicellulases. In order to improve protein production of T. reesei the transcriptional regulation of cellulases and secretory pathway factors have been extensively studied. However, the metabolism of T. reesei under protein production conditions has not received much attention. RESULTS: To understand the physiology and metabolism of T. reesei under protein production conditions we carried out a well-controlled bioreactor experiment with extensive analysis. We used minimal media to make the data amenable for modelling and three strain pairs to cover different protein production levels. With RNA-sequencing transcriptomics we detected the concentration of the carbon source as the most important determinant of the transcriptome. As the major transcriptional response concomitant to protein production we detected the induction of selected genes that were putatively regulated by xyr1 and were related to protein transport, amino acid metabolism and transcriptional regulation. We found novel metabolic responses such as production of glycerol and a cellotriose-like compound. We then used this cultivation data for flux balance analysis of T. reesei metabolism and demonstrate for the first time the use of genome wide stoichiometric metabolic modelling for T. reesei. We show that our model can predict protein production rate and provides novel insight into the metabolism of protein production. We also provide this unprecedented cultivation and transcriptomics data set for future modelling efforts. CONCLUSIONS: The use of stoichiometric modelling can open a novel path for the improvement of protein production in T. reesei. Based on this we propose sulphur assimilation as a major limiting factor of protein production. As an organism with exceptional protein production capabilities modelling of T. reesei can provide novel insight also to other less productive organisms.

Whole-genome metabolic model of Trichoderma reesei built by comparative reconstruction.

BACKGROUND: Trichoderma reesei is one of the main sources of biomass-hydrolyzing enzymes for the biotechnology industry. There is a need for improving its enzyme production efficiency. The use of metabolic modeling for the simulation and prediction of this organism's metabolism is potentially a valuable tool for improving its capabilities. An accurate metabolic model is needed to perform metabolic modeling analysis. RESULTS: A whole-genome metabolic model of T. reesei has been reconstructed together with metabolic models of 55 related species using the metabolic model reconstruction algorithm CoReCo. The previously published CoReCo method has been improved to obtain better quality models. The main improvements are the creation of a unified database of reactions and compounds and the use of reaction directions as constraints in the gap-filling step of the algorithm. In addition, the biomass composition of T. reesei has been measured experimentally to build and include a specific biomass equation in the model. CONCLUSIONS: The improvements presented in this work on the CoReCo pipeline for metabolic model reconstruction resulted in higher-quality metabolic models compared with previous versions. A metabolic model of T. reesei has been created and is publicly available in the BIOMODELS database. The model contains a biomass equation, reaction boundaries and uptake/export reactions which make it ready for simulation. To validate the model, we dem1onstrate that the model is able to predict biomass production accurately and no stoichiometrically infeasible yields are detected. The new T. reesei model is ready to be used for simulations of protein production processes.

Imbalance of plasma amino acids, metabolites and lipids in patients with lysinuric protein intolerance (LPI).

BACKGROUND: Lysinuric protein intolerance (LPI [MIM 222700]) is an aminoaciduria with defective transport of cationic amino acids in epithelial cells in the small intestine and proximal kidney tubules due to mutations in the SLC7A7 gene. LPI is characterized by protein malnutrition, failure to thrive and hyperammonemia. Many patients also suffer from combined hyperlipidemia and chronic kidney disease (CKD) with an unknown etiology. METHODS: Here, we studied the plasma metabolomes of the Finnish LPI patients (n=26) and healthy control individuals (n=19) using a targeted platform for analysis of amino acids as well as two analytical platforms with comprehensive coverage of molecular lipids and polar metabolites. RESULTS: Our results demonstrated that LPI patients have a dichotomy of amino acid profiles, with both decreased essential and increased non-essential amino acids. Altered levels of metabolites participating in pathways such as sugar, energy, amino acid and lipid metabolism were observed. Furthermore, of these metabolites, myo-inositol, threonic acid, 2,5-furandicarboxylic acid, galactaric acid, 4-hydroxyphenylacetic acid, indole-3-acetic acid and beta-aminoisobutyric acid associated significantly (P<0.001) with the CKD status. Lipid analysis showed reduced levels of phosphatidylcholines and elevated levels of triacylglycerols, of which long-chain triacylglycerols associated (P<0.01) with CKD. CONCLUSIONS: This study revealed an amino acid imbalance affecting the basic cellular metabolism, disturbances in plasma lipid composition suggesting hepatic steatosis and fibrosis and novel metabolites correlating with CKD in LPI. In addition, the CKD-associated metabolite profile along with increased nitrite plasma levels suggests that LPI may be characterized by increased oxidative stress and apoptosis, altered microbial metabolism in the intestine and uremic toxicity.

Serum Lipid and Serum Metabolite Components in relation to anthropometric parameters in EPIC-Potsdam participants.

BACKGROUND/AIM: Lipidomic and metabolomic techniques become more and more important in human health research. Recent developments in analytical techniques enable the investigation of high amounts of substances. The high numbers of metabolites and lipids that are detected with among others mass spectrometric techniques challenge in most cases the statistical processes to bring out stable and interpretable results. This study targets to use the novel non-established statistical method treelet transform (TT) to investigate high numbers of metabolites and lipids and to compare the results with the established method principal component analysis (PCA). Serum lipid and metabolite profiles are investigated regarding their association to anthropometric parameters associated to obesity. METHODS: From 226 participants of the EPIC (European Prospective Investigation into Cancer and Nutrition)-Potsdam study blood samples were investigated with an untargeted metabolomics approach regarding serum metabolites and lipids. Additionally, participants were surveyed anthropometrically to assess parameters of obesity, such as body mass index (BMI), waist-to-hip-ratio (WHR) and body fat mass. TT and PCA are used to generate treelet components (TCs) and factors summarizing serum metabolites and lipids in new, latent variables without too much loss of information. With partial correlations TCs and factors were associated to anthropometry under the control for relevant parameters, such as sex and age. RESULTS: TT with metabolite variables (p=121) resulted in 5 stable and interpretable TCs explaining 18.9% of the variance within the data. PCA on the same variables generated 4 quite complex, less easily interpretable factors explaining 37.5% of the variance. TT on lipidomic data (p=353) produced 3 TCs as well as PCA on the same data resulted in 3 factors; the proportion of explained variance was 17.8% for TT and 39.8% for PCA. In both investigations TT ended up with stable components that are easier to interpret than the factors from the PCA. In general, the generated TCs and factors were similar in their structure when the factors are considered regarding the original variables loading high on them. Both TCs and factors showed associations to anthropometric measures. CONCLUSIONS: TT is a suitable statistical method to generate summarizing, latent variables in data sets with more variables than observations. In the present investigation it resulted in similar latent variables compared to the established method of PCA. Whereby less variance is explained by the summarizing constructs of TT compared to the factors of PCA, TCs are easier to interpret. Additionally the resulting TCs are quite stable in bootstrap samples.

Increased dihydroceramide/ceramide ratio mediated by defective expression of degs1 impairs adipocyte differentiation and function.

Adipose tissue dysfunction is an important determinant of obesity-associated, lipid-induced metabolic complications. Ceramides are well-known mediators of lipid-induced insulin resistance in peripheral organs such as muscle. DEGS1 is the desaturase catalyzing the last step in the main ceramide biosynthetic pathway. Functional suppression of DEGS1 activity results in substantial changes in ceramide species likely to affect fundamental biological functions such as oxidative stress, cell survival, and proliferation. Here, we show that degs1 expression is specifically decreased in the adipose tissue of obese patients and murine models of genetic and nutritional obesity. Moreover, loss-of-function experiments using pharmacological or genetic ablation of DEGS1 in preadipocytes prevented adipogenesis and decreased lipid accumulation. This was associated with elevated oxidative stress, cellular death, and blockage of the cell cycle. These effects were coupled with increased dihydroceramide content. Finally, we validated in vivo that pharmacological inhibition of DEGS1 impairs adipocyte differentiation. These data identify DEGS1 as a new potential target to restore adipose tissue function and prevent obesity-associated metabolic disturbances.

Overexpression of PPARγ specifically in pancreatic ß-cells exacerbates obesity-induced glucose intolerance, reduces ß-cell mass, and alters islet lipid metabolism in male mice.

The contribution of peroxisomal proliferator-activated receptor (PPAR)-γ agonism in pancreatic ß-cells to the antidiabetic actions of thiazolidinediones has not been clearly elucidated. Genetic models of pancreatic ß-cell PPARγ ablation have revealed a potential role for PPARγ in ß-cell expansion in obesity but a limited role in normal ß-cell physiology. Here we overexpressed PPARγ1 or PPARγ2 specifically in pancreatic ß-cells of mice subjected to high-fat feeding using an associated adenovirus (ß-PPARγ1-HFD and ß-PPARγ2-HFD mice). We show ß-cell-specific PPARγ1 or PPARγ2 overexpression in diet-induced obese mice exacerbated obesity-induced glucose intolerance with decreased ß-cell mass, increased islet cell apoptosis, and decreased plasma insulin compared with obese control mice (ß-eGFP-HFD mice). Analysis of islet lipid composition in ß-PPARγ2-HFD mice revealed no significant changes in islet triglyceride content and an increase in only one of eight ceramide species measured. Interestingly ß-PPARγ2-HFD islets had significantly lower levels of lysophosphatidylcholines, lipid species shown to enhance insulin secretion in ß-cells. Gene expression profiling revealed increased expression of uncoupling protein 2 and genes involved in fatty acid transport and ß-oxidation. In summary, transgenic overexpression of PPARγ in ß-cells in diet-induced obesity negatively impacts whole-animal carbohydrate metabolism associated with altered islet lipid content, increased expression of ß-oxidative genes, and reduced ß-cell mass.

Steroid and steroid glucuronide profiles in urine during pregnancy determined by liquid chromatography-electrospray ionization-tandem mass spectrometry.

An ultra performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-MS/MS) method was developed for the analysis of steroids and their glucuronides in urine samples. The method provides high sensitivity and fast analysis, as both steroids and their glucuronides can be analyzed directly without hydrolysis or complex sample preparation. The method was applied in profiling of targeted and nontargeted steroids and steroid glucuronides during pregnancy. The concentrations of 11 of 27 targeted steroids and steroid glucuronides and the concentrations of 25 nontargeted steroid glucuronides increased about 10-400 fold during the pregnancy. The concentrations of most of these 36 compounds began to increase in the first days of the pregnancy, increased gradually during the pregnancy, achieved a maximum in late pregnancy, and decreased sharply after delivery. Exceptionally, the concentrations of allopregnanolone and 17-hydroxypregnenolone started to increase later than those of the other steroids. Moreover, the concentrations of E2 glucuronides began to decrease one week before the delivery, in contrast to most of the steroids and steroid glucuronides, whose concentrations dropped sharply during the delivery. Concentrations of 34 compounds decreased noticeably when the subject was on sick leave owing a series of painful contractions. The results suggest that steroids and especially steroid glucuronides may provide a valuable diagnostic tool to follow the course of pregnancy.

Cord serum lipidome in prediction of islet autoimmunity and type 1 diabetes.

Previous studies show that children who later progress to type 1 diabetes (T1D) have decreased preautoimmune concentrations of multiple phospholipids as compared with nonprogressors. It is still unclear whether these changes associate with development of ß-cell autoimmunity or specifically with clinical T1D. Here, we studied umbilical cord serum lipidome in infants who later developed T1D (N = 33); infants who developed three or four (N = 31) islet autoantibodies, two (N = 31) islet autoantibodies, or one (N = 48) islet autoantibody during the follow-up; and controls (N = 143) matched for sex, HLA-DQB1 genotype, city of birth, and period of birth. The analyses of serum molecular lipids were performed using the established lipidomics platform based on ultra-performance liquid chromatography coupled to mass spectrometry. We found that T1D progressors are characterized by a distinct cord blood lipidomic profile that includes reduced major choline-containing phospholipids, including sphingomyelins and phosphatidylcholines. A molecular signature was developed comprising seven lipids that predicted high risk for progression to T1D with an odds ratio of 5.94 (95% CI, 1.07-17.50). Reduction in choline-containing phospholipids in cord blood therefore is specifically associated with progression to T1D but not with development of ß-cell autoimmunity in general.