Omics insights into extracellular vesicles in embryo implantation and their therapeutic utility

Implantation success relies on intricate interplay between the developing embryo and the maternal endometrium. Extracellular vesicles (EVs) represent an important player of this intercellular signalling through delivery of functional cargo (proteins and RNAs) that reprogram the target cells protein and RNA landscape. Functionally, the signalling reciprocity of endometrial and embryo EVs regulates the site of implantation, preimplantation embryo development and hatching, antioxidative activity, embryo attachment, trophoblast invasion, arterial remodelling, and immune tolerance. Omics technologies including mass spectrometry have been instrumental in dissecting EV cargo that regulate these processes as well as molecular changes in embryo and endometrium to facilitate implantation. This has also led to discovery of potential cargo in EVs in human uterine fluid (UF) and embryo spent media (ESM) of diagnostic and therapeutic value in implantation success, fertility, and pregnancy outcome. This review discusses the contribution of EVs in functional hallmarks of embryo implantation, and how the integration of various omics technologies is enabling design of EV-based diagnostic and therapeutic platforms in reproductive medicine.     

Insights into cancer and neurodegenerative diseases through selenoproteins and the connection with gut microbiota – current analytical methodologies

ABSTRACT

Introduction: Selenium plays many key roles in health especially in connection with cancer and neurodegenerative diseases. However, it needs to be appreciated that the essentiality/toxicity of selenium depends on both, a narrow range of concentration and the chemical specie involved. In this context, selenoproteins are essential biomolecules against these disorders, mainly due to its antioxidant action. To this end, analytical methodologies may allow identifying and quantifying individual selenospecies in human biofluids and tissues.

Areas covered: This review focus on the role of selenoproteins in medicine, with special emphasis in cancer and neurodegenerative diseases, considering the possible link with gut microbiota. In particular, this article reviews the analytical techniques and procedures recently developed for the absolute quantification of selenoproteins and selenometabolites in human biofluids and tissues.

Expert commentary: The beneficial role of selenium in human health has been extensively studied and reviewed. However, several challenges remain unsolved as discussed in this article: (i) speciation of selenium (especially selenoproteins) in cancer and neurodegenerative disease patients; (ii) supplementation of selenium in humans using functional foods and nutraceuticals; (iii) the link between selenium and selenoproteins expression and the gut microbiota and (iv) analytical methods and pitfalls for the absolute quantification of selenoproteins and selenometabolites.     

Regulation of cellular immune responses by selenium

Selenium (Se) is an essential nutritional factor that affects the development and expression of cell-mediated immune responses directed toward malignant cells. These studies have shown that dietary (2 ppm for 8 wk) or in in vitro (1×10⁻⁷ M) supplementation with Se (as sodium selenite) results in a significant enhancement of the proliferative responses of spleen lymphocytes from C57B1/6J mice in response to stimulation with mitogen or antigen. Se deficiency (0.02 ppm for 8 wk) had the opposite effect. The alterations in the ability of the cells to proliferate, which occurred in the absence of changes in the endogenous levels of interleukin-2 (II₂) or interleukin 1, were apparently related to the ability of Se to alter the kinetics of expression of high-affinity Il₂ receptors on the surface of activated lymphocytes. This resulted in an enhanced or delayed clonal expansion of the cells, and in an increased or decreased frequency of cytotoxic cells within a given cell population. The changes in tumor cytotoxicity were paralleled by changes in the amounts of lymphotoxin produced by the activated cells. Dietary Se modulations had a comparable effect on macrophage-mediated tumor cytodestruction. The results also suggested that Se exerts its effect 8–24 h after stimulation, and that it most likely affects processes in the cytoplasmic and/or nuclear compartments of activated lymphocytes.     

New insights into cardiovascular and lipid metabolomics

Metabolomics is the study of metabolite profiles in biological samples, particularly urine, saliva, blood plasma and fat biopsies. The metabolome is now considered by some to be the most predictive phenotype: consequently, the comprehensive and quantitative study of metabolites is a desirable tool for diagnosing disease, identifying new therapeutic targets and enabling appropriate treatments. A wealth of information about metabolites has been accumulated with global profiling tools and several candidate technologies for metabolomic studies are now available. Many high-throughput metabolomics methodologies are currently under development and have yet to be applied in clinical practice on a routine basis. In the cardiovascular field, few recent metabolomic studies have been reported so far. This minireview provides an updated overview of alternative technical approaches for metabolomics studies and reviews initial applications of metabolomics that relate to both cardiovascular disease and lipid metabolism research.     

A Multi‐Omics Analysis of Glycine max Leaves Reveals Alteration in Flavonoid and Isoflavonoid Metabolism Upon Ethylene and Abscisic Acid Treatment

Phytohormones are central to plant growth and development. Despite the advancement in our knowledge of hormone signaling, downstream targets, and their interactions upon hormones action remain largely fragmented, especially at the protein and metabolite levels. With an aim to get new insight into the effects of two hormones, ethylene (ET) and abscisic acid (ABA), this study utilizes an integrated proteomics and metabolomics approach to investigate their individual and combined (ABA+ET) signaling in soybean leaves. Targeting low‐abundance proteins, our previously established protamine sulfate precipitation method was applied, followed by label‐free quantification of identified proteins. A total of 4129 unique protein groups including 1083 differentially modulated in one (individual) or other (combined) treatments were discerned. Functional annotation of the identified proteins showed an increased abundance of proteins related to the flavonoid and isoflavonoid biosynthesis and MAPK signaling pathway in response to ET treatment. HPLC analysis showed an accumulation of isoflavones (genistin, daidzein, and genistein) upon ET treatment, in agreement with the proteomics results. A metabolome analysis assigned 79 metabolites and further confirmed the accumulation of flavonoids and isoflavonoids in response to ET. A potential cross‐talk between ET and MAPK signaling, leading to the accumulation of flavonoids and isoflavonoids in soybean leaves is suggested.     

Omics technologies provide new insights into the molecular physiopathology of equine osteochondrosis

Background

Osteochondrosis (OC(D)) is a juvenile osteo-articular disorder affecting several mammalian species. In horses, OC(D) is considered as a multifactorial disease and has been described as a focal disruption of endochondral ossification leading to the development of osteoarticular lesions. Nevertheless, OC(D) physiopathology is poorly understood. Affected horses may present joint swelling, stiffness and lameness. Thus, OC(D) is a major concern for the equine industry. Our study was designed as an integrative approach using omics technologies for the identification of constitutive defects in epiphyseal cartilage and/or subchondral bone associated with the development of primary lesions to further understand OC(D) pathology. This study compared samples from non-affected joints (hence lesion-free) from OC(D)-affected foals (n = 5, considered predisposed samples) with samples from OC-free foals (n = 5) considered as control samples. Consequently, results are not confounded by changes associated with the evolution of the lesion, but focus on altered constitutive molecular mechanisms. Comparative proteomics and micro computed tomography analyses were performed on predisposed and OC-free bone and cartilage samples. Metabolomics was also performed on synovial fluid from OC-free, OC(D)-affected and predisposed joints.

Results

Two lesion subtypes were identified: OCD (lesion with fragment) and OC (osteochondral defects). Modulated proteins were identified using omics technologies (2-DE proteomics) in cartilage and bone from affected foals compare to OC-free foals. These were associated with cellular processes including cell cycle, energy production, cell signaling and adhesion as well as tissue-specific processes such as chondrocyte maturation, extracellular matrix and mineral metabolism. Of these, five had already been identified in synovial fluid of OC-affected foals: ACTG1 (actin, gamma 1), albumin, haptoglobin, FBG (fibrinogen beta chain) and C4BPA (complement component 4 binding protein, alpha).

Conclusion

This study suggests that OCD lesions may result from a cartilage defect whereas OC lesions may be triggered by both bone and cartilage defects, suggesting that different molecular mechanisms responsible for the equine osteochondrosis lesion subtypes and predisposition could be due to a defect in both bone and cartilage. This study will contribute to refining the definition of OC(D) lesions and may improve diagnosis and development of therapies for horses and other species, including humans.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-947) contains supplementary material, which is available to authorized users.     

Comparative proteomic analysis provides new insights into mulberry dwarf responses in mulberry (Morus alba L.)

Mulberry dwarf (MD) is a serious infectious disease of mulberry caused by phytoplasma. Infection with MD phytoplasma results in stress phenotypes of yellowing, phyllody, stunting, proliferation, and witches' broom. Physiological and biochemical analysis has shown that infection with MD phytoplasma causes an increase in soluble carbohydrate and starch content, and a decrease in the net photosynthesis rate, carboxylation efficiency, and pigment content of leaves. Furthermore, damage to the chloroplast ultrastructure was detected in infected leaves. To better understand the pathogen‐stress response of mulberry (Morus alba L.) to MD phytoplasma, we conducted a comparative proteomic analysis using 2‐DE of infected and healthy leaves. Among 500 protein spots that were reproducibly detected, 20 were down‐regulated and 17 were up‐regulated. MS identified 16 differentially expressed proteins. The photosynthetic proteins rubisco large subunit, rubisco activase, and sedoheptulose‐1,7‐bisphosphatase showed enhanced degradation in infected leaves. Based these results, a model for the occurrence mechanism of MD is proposed. In conclusion, this study provides new insights into the mulberry response to MD phytoplasma infection.     

Different bioavailability in humans of wheat and fish selenium as measured by blood platelet response to increased dietary Se

The bioavailabilities of selenium (Se) from Se-rich fish species and Se-rich wheat were compared in a study involving 32 healthy volunteers. Initial serum Se values were 109±16 μg/L (mean±SD). For 6 wk, one group (n=11) included Se-rich bread in their diet, bringing daily average intake of Se up to 135±25 μg/d. Another group (n=11) consumed Se-rich fish daily (average Se intake: 115±31 μg/d), whereas the control group (n=10) ate their normal diet, providing 77±25 μg Se/d. Serum Se increased by 17% (P<0.01), and platelet Se increased by 30% (P<0.01) in the wheat group. Although platelet Se decreased by 11% in the fish group, no changes in serum and platelet Se in the fish or control group reached statistical significance. Glutathione peroxidase (EC 1.11.1.9; GSH-Px) activity in serum and platelets did not change during the study, nor did platelet mercury (Hg) content. Since the dietary intake of Hg, arsenium (As), and fatty acids could not satisfactorily explain the lack of response in the fish group, the results are indicative of low bioavailability of fish Se in humans. At present, wheat Se seems to be the most important factor contributing to the body stores of Se in this study population. Dr. Norheim died on January 9, 1991.     

Proteomic insights into seed germination in response to environmental factors

Seed germination is a critical process in the life cycle of higher plants. During germination, the imbibed mature seed is highly sensitive to different environmental factors. However, knowledge about the molecular and physiological mechanisms underlying the environmental effects on germination has been lacking. Recent proteomic work has provided invaluable insight into the molecular processes in germinating seeds of Arabidopsis, rice (Oryza sativa), soybean (Glycine max), barley (Hordeum vulgare), maize (Zea mays), tea (Camellia sinensis), European beech (Fagus sylvatica), and Norway maple (Acer platanoides) under different treatments including metal ions (e.g. copper and cadmium), drought, low temperature, hormones, and chemicals (gibberellic acid, abscisic acid, salicylic acid, and α‐amanitin), as well as Fusarium graminearum infection. A total of 561 environmental factor‐responsive proteins have been identified with various expression patterns in germinating seeds. The data highlight diverse regulatory and metabolic mechanisms upon seed germination, including induction of environmental factor‐responsive signaling pathways, seed storage reserve mobilization and utilization, enhancement of DNA repair and modification, regulation of gene expression and protein synthesis, modulation of cell structure, and cell defense. In this review, we summarize the interesting findings and discuss the relevance and significance for our understanding of environmental regulation of seed germination.     

Integrated analysis of proteomics-delineated and metabolomics-delineated hepatic metabolic responses to (−)-hydroxycitric acid in chick embryos

(−)-Hydroxycitric acid [(−)-HCA] is widely used as a nutritional supplement to control body weight and fat accumulation in animals and humans, whereas the underlying biochemical mechanism is unclear. Broiler chicken was used as a model for studies of obesity due to its natural hyperglycemia and being insulin resistant. The current study aimed to obtain a systematic view of serum metabolites and hepatic proteins and well understand the mechanism of hepatic metabolic response to (−)-HCA treatment in chick embryos. The results showed that 22, 90, and 82 of differentially expressed proteins were identified at E14d, E19d, and H1d in chick embryos treated with (−)-HCA, respectively. Meanwhile, 5, 83, and 88 of serum metabolites significantly changed at E14d, E19d, and H1d in chick embryos after (−)-HCA treatment. Bioinformatics analysis showed that the key proteins and metabolites, which were significantly altered in chick embryos treated with (−)-HCA, were mainly involved in the citrate cycle, glycolysis/gluconeogenesis, fatty acid metabolism, and pyruvate metabolism. Our data indicated that (−)-HCA treatment might promote fat metabolism via regulating the key protein expression levels and metabolite contents in the citrate cycle, glycolysis/gluconeogenesis, and oxidative phosphorylation during chicken embryonic development. These results will deepen our understanding of the mechanism of fat reduction by (−)-HCA and provide substantial information for (−)-HCA as a nutritional supplement to control body weight gain and curb obesity-related diseases.     

Effects of dietary selenium supplementation on tissue selenium distribution and glutathione peroxidase activity in Chinese Ring Necked Pheasants

The objective of this study was to determine the concentration of total selenium (Se) and the proportions of total Se comprised as selenomethionine (SeMet) and selenocysteine (SeCys) in the postmortem tissues of female pheasants (Phasianus Colchicus Torquator) offered diets that contained graded additions of selenised-enriched yeast (SY) or a single comparative dose of sodium selenite (SS). Thiobarbituric acid reactive substances (TBARS) and tissue glutathione peroxidase (GSH-Px) activity of breast (Pectoralis Major) were assessed at 0 and 5 days postmortem. A total of 216 female pheasant chicks were enrolled into the study. Twenty-four birds were euthanased at the start of the study, and samples of blood, breast muscle, leg muscle (M. Peroneus Longus and M. Gastrocnemius), heart, liver, kidney and gizzard were collected for determination of total Se. Remaining birds were blocked by live weight and randomly allocated to one of four dietary treatments (n = 48 birds/treatment) that either differed in Se source (SY v. SS) or dose (control (0.17 mg total Se/kg), SY-L and SS-L (0.3 mg/kg total Se as SY and SS, respectively) and SY-H (0.45 mg total Se/kg)). Following 42 and 91 days of treatment, 24 birds per treatment were euthanased, and samples of blood, breast muscle, leg muscle, heart, liver, kidney and gizzard were retained for determination of total Se and the proportion of total Se comprised as SeMet or SeCys. Whole blood GSH-Px activity was determined at each time point. Tissue GSH-Px activity and TBARS were determined in breast tissue at the end of the study. There were increases in both blood and tissues to the graded addition of SY to the diet (P < 0.001), but the same responses were not apparent with the blood and tissues of selenite-supplemented birds receiving a comparable dose (SY-L v. SS-L). Although there were differences between tissue types in the distribution of SeMet and SeCys, there were few differences between treatments. There were effects of treatment on erythrocyte GSH-Px activity (P = 0.012) with values being higher in treatments SY-H and SS-L when compared with the negative control and treatment SY-L. There were no effects of treatment on tissue GSH-Px activity, which is reflected in the overall lack of any treatment effects on TBARS.     

Stream ecosystem response to, and recovery from, experimental exposure to selenium

The effects of selenium on streamecosystems were studied in outdoor,experimental stream mesocosms during a dosingperiod in which sodium selenite was added atnominal concentrations of 30 µg/L,10 µg/L, and 2.5 µg/L. The durationof the high, medium, and low treatments were573 d, 972 d, and 311 d, respectively. Apost-dosing period of three years (hightreatment) and two years (medium, lowtreatments) also was studied. Seleniumconcentrations in water, sediment, plants, andmacroinvertebrates were measured throughoutthe dosing and recovery periods. Fatheadminnows and bluegill sunfish were periodicallyheld in the streams to measure seleniumaccumulation and its effects on fish survivaland reproduction. Quantitative samples ofmacroinvertebrates were collected to assessselenium effects on macroinvertebratecommunities.Mean selenium concentration inwater was quite close to the nominalconcentration. Selenium accumulated in thesediment in all three treated streams, but notin the control streams. Sediment seleniumdecreased slowly after dosing ceased, but wasstill significantly higher than in controlstreams three years (high treatment) and twoyears (medium treatment) later.Macrophytetissue selenium concentrations weresignificantly greater in all three treatmentsthan those in the control streams duringdosing. Macrophyte selenium bioaccumulationfactors (BAFs) ranged from about 300 to 1900. Tissue selenium decreased rapidly in all threetreatments after dosing ended.During dosing,selenium concentrations in animals from allthree treatments were significantly higherthan in those from control streams. The BAFsfor macroinvertebrates ranged from 1100 to2000. Isopods accumulated more, and amphipodsless, selenium than other invertebrates. Therewere no significant effects of selenium onmacroinvertebrate abundance, richness ordiversity. Several macroinvertebrates werenot affected by exposure to selenium, butisopod and Tubifex populations weredramatically reduced in the high and mediumtreatments. After dosing, mean seleniumconcentration in macroinvertebrates decreasedslowly.Bluegill sunfish accumulated seleniumduring dosing and after selenium additionsceased. Tissue selenium was highest in theliver, followed by the gonads, skeletalmuscle, and whole body. Tissue seleniumconcentrations one (high, medium) and two(high) years after dosing were lower thanduring dosing, but whole body, skeletal muscleand liver concentrations were high enough tobe considered potentially toxic.Recovery ofselenium contaminated streams includes bothreduction of tissue selenium concentration tonon-toxic levels in fish and their foodorganisms and recovery of populations of taxadeleteriously affected by selenium exposure. Our results suggest that when selenium iseliminated from the water in streams, seleniumconcentrations in sediment, plants,macroinvertebrates, and fishes will decreaseto levels that approach concentrationsconsidered to be non-toxic to fish andwildlife and that affected populations willrecover within several years. Based onselenium accumulation in the food chain andthe presence of real, but not statisticallysignificant, effects on fish mortality andreproduction in the low treatment streams, wesupport a selenium water quality criterion forthe protection of fishes and sensitiveinvertebrates of 2 µg/L or less.     

Characterization and expression analysis of seven selenoprotein genes in yellow catfish Pelteobagrus fulvidraco to dietary selenium levels

BackgroundSelenium (Se) appears in the selenoproteins in the form of selenocysteine (Sec) and is important for the growth and development of vertebrates. The present study characterized seven selenoproteins, consisting of the GPX1, GPX3, GPX4, SELENOW, SELENOP, TXNRD2 and TXNRD3 cDNAs in various tissues of yellow catfish, explored their regulation to dietary Se addition.Methods3′ and 5′ RACE PCR were used to clone full-length cDNA sequences of seven selenoprotein genes (GPX1, GPX3, GPX4, SELENOW, SELENOP, TXNRD2 and TXNRD3). Their molecular characterizations were analyzed, including conservative motifs and the SECIS elements. The phylogenetic trees were generated through neighbor-joining (NJ) method with MEGA 6.0 with 1000 bootstrap replications. Quantitative real-time PCR was used to explore their mRNA tissue distribution in the heart, anterior intestine, dorsal muscle, head kidney, gill, liver, brain, spleen and mesenteric fat. Yellow catfish (mixed sex) were fed diets with dietary Se contents at 0.03 (low Se), 0.25 (adequate Se) and 6.39 (high Se) mg Se/kg, respectively, for 12 weeks, and their spleen, kidney, testis and brain were used for the determination of the mRNA levels of the seven selenoproteins.ResultsThe seven selenoproteins had similar domains to their corresponding members of other vertebrates. They were widely expressed in nine tissues, including heart, liver, brain, spleen, head kidney, dorsal muscle, mesenteric fat, anterior intestine and gill, but showed tissue-dependent expression patterns. Dietary Se addition affected the expression of the seven genes in spleen, kidney, testis and brain tissues of yellow catfish.ConclusionTaken together, our study demonstrated the characterization, expression and regulation of seven selenoproteins, which increased our understanding of the biological functions of Se and selenoproteins in fish.     

New insights into the placebo and nocebo responses

In modern medicine, the placebo response or placebo effect has often been regarded as a nuisance in basic research and particularly in clinical research. The latest scientific evidence has demonstrated, however, that the placebo effect and the nocebo effect, the negative effects of placebo, stem from highly active processes in the brain that are mediated by psychological mechanisms such as expectation and conditioning. These processes have been described in some detail for many diseases and treatments, and we now know that they can represent both strength and vulnerability in the course of a disease as well as in the response to a therapy. However, recent research and current knowledge raise several issues that we shall address in this review. We will discuss current neurobiological models like expectation-induced activation of the brain reward circuitry, Pavlovian conditioning, and anxiety mechanisms of the nocebo response. We will further explore the nature of the placebo responses in clinical trials and address major questions for future research such as the relationship between expectations and conditioning in placebo effects, the existence of a consistent brain network for all placebo effects, the role of gender in placebo effects, and the impact of getting drug-like effects without drugs.