Key Biochemical Pathways Research Articles

Impact of the long-term storage on flavor quality of Liupao tea using sensory evaluation combined with metabolomics analysis

This study comprehensively investigated the impact of different storage times on the quality and metabolomic profiles of Liupao tea (LPT). The sensory evaluations revealed that both Maosheng (MS) and Tianyu (TY) teas exhibited a browning of tea appearance and brightening of tea infusion during storage. The taste evolved from bitterness and astringency to purity and briskness, while the aroma shifted from stuffy to woody and aged aromas. Notably, MS teas exhibited superior sensory quality after 10 years, while TY teas reached optimal quality in the 8th year of storage. Correlation analysis of metabolites and sensory attributes has underscored the integral influence of metabolites throughout the storage process, which significantly directed the development of tea quality. The non-volatile metabolites exerted significant influence on tea flavor by modulating key biochemical pathways, including the oxidation of catechins, the formation of alkaloids as well as the glycosylation and/or methylation of flavonoids. However, TY teas experienced both glycosylation and methylation, which promoted the transformation of bitterness and astringency, achieving a mellow and brisk taste more quickly than MS teas. The transformation pathways of volatile metabolites potentially involved the hydrolysis of linalool glycosides and phenylethanol glycosides, the synthesis of sesquiterpenes, the methylation of gallic acid and the degradation of carotenoids. However, the divergent trends observed in ketones and aldehydes between the two types of tea could culminate in distinct aromatic profiles, which might be due to different metabolic pathways or differences in the rates of metabolite formation and degradation during storage. Additionally, the antioxidant analysis revealed that both MS and TY teas exhibited a parabolic trend in comprehensive antioxidant capacity during storage, which primarily influenced by the oxidative polymerization of phenolic compounds and the glycosylation of flavonoids. In summary, this study emphasized the multifaceted attributes of tea quality and the importance of metabolites in shaping sensory quality and health properties. It was found that the optimal storage time of 8 to 10 years for LPT was conducive to attaining a desirable balance of flavor and health benefits.

Hyperexcitability and translational phenotypes in a preclinical mouse model of SYNGAP1-related intellectual disability

Disruption of SYNGAP1 directly causes a genetically identifiable neurodevelopmental disorder (NDD) called SYNGAP1-related intellectual disability (SRID). Without functional SynGAP1 protein, individuals are developmentally delayed and have prominent features of intellectual disability (ID), motor impairments, and epilepsy. Over the past two decades, there have been numerous discoveries indicating the critical role of Syngap1. Several rodent models with a loss of Syngap1 have been engineered, identifying precise roles in neuronal structure and function, as well as key biochemical pathways key for synapse integrity. Homozygous loss of SYNGAP1/Syngap1 is lethal. Heterozygous mutations of Syngap1 result in a broad range of behavioral phenotypes. Our in vivo functional data, using the original mouse model from the Huganir laboratory, corroborated behaviors including robust hyperactivity and deficits in learning and memory in young adults. Furthermore, we described impairments in the domain of sleep, characterized using neurophysiological data that was collected with wireless, telemetric electroencephalography (EEG). Syngap1+/− mice exhibited elevated spiking events and spike trains, in addition to elevated power, most notably in the delta power frequency. For the first time, we illustrated that primary neurons from Syngap1+/− mice displayed: 1) increased network firing activity, 2) greater bursts, 3) and shorter inter-burst intervals between peaks, by utilizing high density microelectrode arrays (HD-MEA). Our work bridges in vitro electrophysiological neuronal activity and function with in vivo neurophysiological brain activity and function. These data elucidate quantitative, translational biomarkers in vivo and in vitro that can be utilized for the development and efficacy assessment of targeted treatments for SRID.

Gestational Long Term High-Altitude Hypoxia and Metabolomic Reprogramming in Uterine Arteries of Adult Sheep

Long term hypoxic stress during pregnancy causes a reduction in uterine blood flow, compromising oxygen and nutrient delivery to the fetus. There are multiple epigenetic factors involved in the structural and functional impairments that underlie the impairments in uterine arterial function. Evidence indicates that oxidative stress plays an important role to the decrements in blood flow following high altitude pregnancy. The purpose of the current study was to test the hypothesis that changes in oxylipin and endocannabinoid signaling are important to the increases in oxidative stress along with changes in glucose and fat metabolism. To further understand the mechanisms associated with disruption of uterine artery blood flow due to high altitude pregnancy, we examined a high altitude pregnant sheep model. Pregnant and non-pregnant sheep were housed either at low (700m) or high altitude (3801 meters) for 110+ days of pregnancy. Uterine arteries were excised from pregnant and non-pregnant sheep and targeted as well as untargeted metabolomic analyses were performed to test the hypothesis that hypoxia increases oxidative stress and reduces oxidative metabolism. Enrichment analysis of the metabolites show that hypoxia impacted several key biochemical pathways. At high altitude there were decreases in Alpha Linolenic Acid and Linoleic Acid Metabolism along with reductions in Pyruvate Metabolism and antioxidants. Further to this, there was an increase in the metabolism of Arachidonic Acid, Glycerolipids, and Methionine, which are linked to inflammation, cell signaling, and cell growth. Fully understanding how increased oxidative stress and inflammatory pathways along with decreased antioxidant capacity and changes in cellular metabolism is tied to malformations in uterine arterial structure and function following high altitude gestation is an active area of investigation. This work was supported by the National Institutes of Health Grants NIH R01HL155295, R01HL149608, R03HD098477, P01HD083132, and U24DK097154. Additional support was provided by the Loma Linda University School of Medicine and West Coast Metabolomics Center. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Nanoplastic toxicity induces metabolic shifts in Populus × euramericana cv. '74/76' revealed by multi-omics analysis

There is increasing global concern regarding the pervasive issue of plastic pollution. We investigated the response of Populus × euramericana cv. ‘74/76’ to nanoplastic toxicity via phenotypic, microanatomical, physiological, transcriptomic, and metabolomic approaches. Polystyrene nanoplastics (PS-NPs) were distributed throughout the test plants after the application of PS-NPs. Nanoplastics principally accumulated in the roots; minimal fractions were translocated to the leaves. In leaves, however, PS-NPs easily penetrated membranes and became concentrated in chloroplasts, causing thylakoid disintegration and chlorophyll degradation. Finally, oxidant damage from the influx of PS-NPs led to diminished photosynthesis, stunted growth, and etiolation and/or wilting. By integrating dual-omics data, we found that plants could counteract mild PS-NP-induced oxidative stress through the antioxidant enzyme system without initiating secondary metabolic defense mechanisms. In contrast, severe PS-NP treatments promoted a shift in metabolic pattern from primary metabolism to secondary metabolic defense mechanisms, an effect that was particularly pronounced during the upregulation of flavonoid biosynthesis. Our findings provide a useful framework from which to further clarify the roles of key biochemical pathways in plant responses to nanoplastic toxicity. Our work also supports the development of effective strategies to mitigate the environmental risks of nanoplastics by biologically immobilizing them in contaminated lands.

Integrating machine learning and nontargeted plasma lipidomics to explore lipid characteristics of premetabolic syndrome and metabolic syndrome.

To identify plasma lipid characteristics associated with premetabolic syndrome (pre-MetS) and metabolic syndrome (MetS) and provide biomarkers through machine learning methods. Plasma lipidomics profiling was conducted using samples from healthy individuals, pre-MetS patients, and MetS patients. Orthogonal partial least squares-discriminant analysis (OPLS-DA) models were employed to identify dysregulated lipids in the comparative groups. Biomarkers were selected using support vector machine recursive feature elimination (SVM-RFE), random forest (rf), and least absolute shrinkage and selection operator (LASSO) regression, and the performance of two biomarker panels was compared across five machine learning models. In the OPLS-DA models, 50 and 89 lipid metabolites were associated with pre-MetS and MetS patients, respectively. Further machine learning identified two sets of plasma metabolites composed of PS(38:3), DG(16:0/18:1), and TG(16:0/14:1/22:6), TG(16:0/18:2/20:4), and TG(14:0/18:2/18:3), which were used as biomarkers for the pre-MetS and MetS discrimination models in this study. In the initial lipidomics analysis of pre-MetS and MetS, we identified relevant lipid features primarily linked to insulin resistance in key biochemical pathways. Biomarker panels composed of lipidomics components can reflect metabolic changes across different stages of MetS, offering valuable insights for the differential diagnosis of pre-MetS and MetS.

Differential voluntary feed intake and whole transcriptome profiling in the hypothalamus of young sheep offered CP and phosphorus-deficient diets.

A reduction in voluntary feed intake is observed in ruminants consuming nutrient-deficient diets, such as those with a low CP or P content, and has been attributed to active metabolic regulation, rather than a physical constraint. The hypothalamus is the key integrator of feed intake regulation in mammals. The objectives of this experiment were to (1) establish a model of metabolic feed intake regulation in ruminants consuming diets of variable CP and P content, and (2) determine key biochemical pathways and influential points of regulation within the hypothalamus. Merino wethers [n=40; 23.7±1.4kg liveweight (mean±SD)] were fed one of five dietary treatments (n=8/treatment) for 63days in individual pens. The treatments included targeted combinations of high (H) and low (L) CP (110 and 55g/kg DM) and high and low P (2.5 and 0.7g/kg DM) with 9MJ metabolisable energy (ME) per kg DM which were fed ad libitum (UMEI; unrestricted ME intake) resulting in four experimental diets (HCP-HP-UMEI, LCP-HP-UMEI, HCP-LP-UMEI and LCP-LP-UMEI). An additional nutritional treatment (HCP-HP-RMEI) restricted intake of the HCP-HP diet to an equivalent ME intake of wethers consuming the LCP-LP-UMEI treatment. Wethers offered the LCP-HP-UMEI, HCP-LP-UMEI and LCP-LP-UMEI treatments consumed 42, 32 and 49% less total DM (P≤0.05), respectively than the HCP-HP-UMEI treatment, and this was not attributable to any physical limitation of the rumen. Plasma concentrations of urea nitrogen and inorganic phosphate indicated that these nutrient deficiencies were successfully established. To assess potential mechanisms, RNA-seq was conducted on samples from the arcuate nucleus (ARC), ventromedial hypothalamus and lateral hypothalamus of the wethers, yielding a total of 301, 8 and 148 differentially expressed genes across all pairwise comparisons, respectively. The expression of NPY, AGRP and CARTPT, known for their regulatory role in mammalian feed intake regulation, had a similar transcriptional response in the ARC of wethers consuming nutrient-deficient treatments and those consuming a ME-restricted treatment, despite these wethers expressing behaviours indicative of satiated and hungry states, respectively. In addition, genes involved with glycolysis (TPI1), the citric acid cycle (CS, OGDH, GLUD1, GOT1) and oxidative phosphorylation (COX5A, ATP5MC1, ATP5F1B, ATP5MC3) were downregulated in the ARC of wethers fed a nutrient deficient (LCP-LP-UMEI) relative to the non-deficient (HCP-HP-UMEI) treatment. In summary, a model for voluntary feed intake restriction was established to determine genome-wide molecular changes in the hypothalamus of young ruminants.

Elucidating the flavour of cooked white asparagus by combining metabolomics and taste panel analysis

A typical feature of cooked white spears of Asparagus officinalis is their specific flavour. This has been the focus of several recent instrumental and panel studies. Few of these report on the comparison of raw and cooked materials which may lead to a deeper understanding of the flavour chemistry and compositional shifts occurring during food preparation. Here, solid-phase microextraction GC-MS and LC-MS analyses of raw and cooked spears were performed to profile asparagus metabolites. A taste panel evaluated flavour attributes of the same materials. Unsupervised and supervised statistical approaches were applied to investigate the profiles and link metabolomics and sensory results. Flavonoids, saponins and medium-chain volatile carbonyls were more abundant in the raw spears while sulphur-containing volatile compounds, oxidized fatty acids and phenolic acids were more abundant in the cooked spears. GC-Olfactometry-MS was also performed and revealed 35 odour-active volatile flavour compounds. We have proposed and discussed the key biochemical pathways and chemical transformations relevant to asparagus ‘flavour’.

Potential of Microalgae as Functional Foods Applied to Mitochondria Protection and Healthy Aging Promotion

The rapid aging of the Western countries’ populations makes increasingly necessary the promotion of healthy lifestyles in order to prevent/delay the onset of age-related diseases. The use of functional foods can significantly help to achieve this aim, thanks to the contribution of biologically active compounds suitable to protect cellular and metabolic homeostasis from damage caused by stress factors. Indeed, the excessive production of reactive oxygen species (ROS), favored by incorrect eating and behavioral habits, are considered causal elements of oxidative stress, which in turn favors tissue and organism aging. Microalgae represent a convenient and suitable functional food because of their extraordinary ability to concentrate various active compounds, comprising omega-3 polyunsaturated fatty acids, sterols, phenolic compounds, carotenoids and others. Within cells, mitochondria are the cellular organelles most affected by the accumulation of molecular damage produced by oxidative stress. Since, in addition to producing the chemical energy for cellular metabolism, mitochondria control numerous cell cycle regulation processes, including intrinsic apoptosis, responses to inflammatory signals and other biochemical pathways, their dysfunction is considered decisive for many pathologies. Among these, some degenerative diseases of the nervous system, cardiovascular system, kidney function and even cancer are found. From this viewpoint, bioactive compounds of microalgae, in addition to possessing high antioxidant properties, can enhance mitochondrial functionality by modulating the expression of numerous protective factors and enzymes, which in turn regulate some essential biochemical pathways for the preservation of the functional integrity of the cell. Here, we summarize the current knowledge on the role played by microalgal compounds in the regulation of the mitochondrial life cycle, expression of protective and reparative enzymes, regulation of intrinsic apoptosis and modulation of some key biochemical pathways. Special attention was paid to the composition of some cultivable microalgae strains selected for their high content of active compounds suitable to protect and improve mitochondrial functions.

Oxidation of hyperpolarized [1-13 C]pyruvate in isolated rat kidneys.

Kidneys play a central role in numerous disorders but current imaging methods have limited utility to probe renal metabolism. Hyperpolarized (HP) 13 C magnetic resonance imaging is uniquely suited to provide metabolite-specific information about key biochemical pathways and it offers the further advantage that renal imaging is practical in humans. This study evaluated the feasibility of hyperpolarization examinations in a widely used model for analysis of renal physiology, the isolated kidney, which enables isolation of renal metabolism from the effects of other organs and validation of HP results by independent measurements. Isolated rat kidneys were supplied with either HP [1-13 C]pyruvate only or HP [1-13 C]pyruvate plus octanoate. Metabolic activity in both groups was confirmed by stable renal oxygen consumption. HP [1-13 C]pyruvate was readily metabolized to [13 C]bicarbonate, [1-13 C]lactate, and [1-13 C]alanine, detectable seconds after HP [1-13 C]pyruvate was injected. Octanoate suppressed but did not eliminate the production of HP [13 C]bicarbonate from [1-13 C]pyruvate. Steady-state flux analyses using non-HP 13 C substrates validated the utilization of HP [1-13 C]pyruvate, as observed by HP 13 C NMR. In the presence of octanoate, lactate is generated from a tricarboxylic acid cycle intermediate, oxaloacetate. The isolated rat kidney may serve as an excellent model for investigating and establishing new HP 13 C metabolic probes for future kidney imaging applications.

The Role of Mineral Deficiencies in Insulin Resistance and Obesity.

Minerals are critical for maintaining overall health. These tiny chemical compounds are responsible for enzymatic activation, maintaining healthy teeth and bones, regulating energy metabolism, enhancing immunity, and aiding muscle and brain function. However, mineral deficiency in the form of inadequate or under nourished intake affects millions of people throughout the world, with well-documented adverse health consequences of malnutrition. Conversely, mineral deficiency may also be a risk factor for Insulin Resistance (IR) and obesity. This review focuses on another, more "less discussed" form of malnutrition, namely mineral deficiency and its contribution to metabolic disorders. At the cellular level, minerals maintain not only molecular communication but also trigger several key biochemical pathways. Disturbances in these processes due to mineral insufficiency may gradually lead to metabolic disorders such as insulin resistance, pre-diabetes, and central obesity, which might lead to renal failure, cardiac arrest, hepatic carcinoma, and various neurodegenerative diseases. Here we discuss the burden of disease promoted by mineral deficiencies and the medical, social, and economic consequences. Mineral deficiency-mediated IR and obesity have a considerable negative impact on individual well-being, physical consideration, and economic productivity. We discuss possible molecular mechanisms of mineral deficiency that may lead to IR and obesity and suggest strategies to counter these metabolic disorders. To protect mankind from mineral nutrient deficiencies, the key is to take a variety of foods in reasonable quantities, such as organic and pasture-raised eggs, low fat dairy, and grass-fed and finished meats, insecticide, and pesticide-free vegetables and fruits.

Proteomic profiling of metabolic proteins as potential biomarkers of radioresponsiveness for colorectal cancer

Surgery, radiation therapy (RT), and chemotherapy are commonly used treatment modalities for CRC management. The locally advanced CRC is managed with preoperative RT or in combination of chemoradiotherapy whereas palliative RT is recommended for metastatic CRC patients to enhance overall survival and reduce distressing symptoms. There are many biomarkers established based on tumour staging, grading and molecular characteristics of patients (e.g., mutation, DNA methylation, and gene expression profiling). Interestingly, none of these markers are adequately validated for RT scheme. In order to establish the radioresponsive biomarker in CRC, we established a mouse xenograft tumour model and applied radiation to the tumours. We identified 9 metabolic proteins, namely PGK1, PGAM1, ENO1, PKM, TKT, GLUD1, LDHA, GAPDH, and MDH2, which are differentially expressed in tumours with different radioresponsiveness. Furthermore, we validated their expression in tumours from the unirradiated, poorly responded and highly responded tumour groups. In addition, we analysed their expressions in clinical samples from the public database. Extensive literature studies shown that these metabolic proteins are associated with key biochemical pathways including, glycolysis, ammonia detoxification, carcinogenesis, and drug responses. Further studies are needed to translate our findings into clinical use. SignificanceWith the increasing incidence of colorectal cancer (CRC) globally, it is crucial to establish strategic treatment protocol by personalizing cancer treatment. Despite the well-established treatment protocols for CRC in the past decades, the mortality remains high. There is a trend of applying personalized treatment to improve patient survival. It has been reported that biomarkers may be used to predict treatment outcomes or to adjust individual treatment protocols. This project aims to identify specific metabolic proteins as biomarkers for CRC radioresponsiveness. Using bioinformatical analysis, we have identified 9 metabolic proteins which could be used as potential biomarkers for radiation therapy in CRC tumours.

Amino acid δ13C fingerprints of nearshore marine autotrophs are consistent across broad spatiotemporal scales: An intercontinental isotopic dataset and likely biochemical drivers

Abstract Carbon isotope fingerprinting, or multivariate analysis using δ13C values of individual compounds, is a powerful tool in ecological studies, particularly measurements of essential amino acids (EAA δ13C). Despite the widespread application of this technique, there has been little methodological validation to determine (a) whether multivariate EAA δ13C signatures (fingerprints) of primary producer groups vary across space and time and (b) what biochemical mechanisms drive these patterns. Here, we evaluate the spatiotemporal consistency in EAA δ13C fingerprints among nearshore primary producers: Chlorophyta (Ulva sp.), Ochrophyta (kelps), particulate organic matter (POM) and phytoplankton, and Rhodophyta. We analysed 135 samples from 14 genera collected in Alaska, California and Chile. The collections included historical museum samples (1896–1980 CE) of the giant kelp, Macrocystis pyrifera. We employed canonical analysis of principal coordinates and generalized linear models (GLMs) to, respectively, characterize isotopic fingerprints and evaluate the effect of taxonomy, local upwelling regimes, ecological setting, and time on individual EAA δ13C values and associated fingerprints. We also calculated amino acid discrimination values (∆13C) to identify key biochemical pathways responsible for these patterns. We found remarkable consistency in EAA δ13C fingerprints of marine algae across space and through time. Kelps and rhodophytes exhibited statistically distinct multivariate isotopic patterns regardless of geographical location, species identity or time (kelps). In contrast, isotopic fingerprints of POM/phytoplankton and Ulva overlapped substantially. GLMs indicated that producer family, presumably due to the presence/absence of carbon concentrating mechanisms, and site locality are important determinants of individual amino acid δ13C values. Taxonomy was also a key variable for EAA δ13C fingerprints. The calculated discrimination values suggest variation in (a) metabolism of pyruvate and oxaloacetate‐derived amino acids and (b) production of storage and structural carbohydrates are responsible for taxonomic differences in isotopic fingerprints. We conclude EAA δ13C fingerprinting is a robust method for tracing the contribution of diverse primary producer taxa to coastal food webs. We show that this technique can be applied to modern and historical samples, as well as consumers collected across continental scales. The high fidelity of EAA δ13C multivariate patterns coupled with biochemical mechanisms provides a powerful framework for future studies of carbon flow across broad biogeographical and ecological contexts. Read the free Plain Language Summary for this article on the Journal blog.

Transcriptome Analysis Revealed the Key Genes and Pathways Involved in Seed Germination of Maize Tolerant to Deep-Sowing.

To improve our understanding of the mechanism of maize seed germination under deep sowing, transcriptome sequencing and physiological metabolism analyses were performed using B73 embryos separated from ungerminated seeds (UG) or seeds germinated for 2 d at a depth of 2 cm (normal sowing, NS) or 20 cm (deep sowing, DS). Gene ontology (GO) analysis indicated that “response to oxidative stress” and “monolayer-surrounded lipid storage body” were the most significant GO terms in up- and down-regulated differentially expressed genes (DEGs) of DS. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis suggested that “phenylpropanoid biosynthesis” and “starch and sucrose metabolism” were critical processes in maize seed germination under deep-sowing conditions. Consistent with DEGs, the activities of superoxide dismutase, catalase, peroxidases and α-amylase, as well as the contents of gibberellin 4, indole acetic acid, zeatin and abscisic acid were significantly increased, while the jasmonic-acid level was dramatically reduced under deep-sowing stress. The expressions of six candidate genes were more significantly upregulated in B73 (deep-sowing-tolerant) than in Mo17 (deep-sowing-sensitive) at 20 cm sowing depth. These findings enrich our knowledge of the key biochemical pathways and genes regulating maize seed germination under deep-sowing conditions, which may help in the breeding of varieties tolerant to deep sowing.

Molecular signatures of aggressive pediatric liver cancer

Liver masses account for 5 to 6% of pediatric cancer, which includes hepatoblastoma (HBL) along with rare cases of hepatocellular carcinoma (HCC). The most dangerous form of pediatric liver cancer is aggressive HBL, which can be characterized by chemo-resistance and multiple nodules or metastases at diagnosis, all correlating with worse clinical prognosis. Despite intensive studies and a significant improvement in overall outcomes, very little is known about the key molecular pathways which determine the aggressiveness of pediatric liver cancer. Although genetic mutations have been reported in aggressive HBL, they represent a low level (1.9% per case) and are found mainly in two genes CTNNB1 and NRF2. Over the past 5 years, our liver biology and tumor group at Cincinnati Children's Hospital Medical Center has investigated molecular signatures of aggressive HBL by examination of fresh tissue specimens, which were studied immediately after surgery to preserve the integrity of key biochemical pathways. Summarization of these high quality HBL samples discovered several critical pathways that are specific for aggressive pediatric liver cancer. These pathways include three characteristics: Conversion of tumor suppressor proteins (TSPs) by posttranslational modifications into oncogenesActivation of specific chromosomal regions, i.e., Aggressive Liver Cancer Domains (ALCDs) within many oncogenes, resulting in increased expression of oncogenesPotential epigenetic mechanisms that open chromatin structure of oncogenes via ALCDs. This commentary summarizes our key findings and discusses development of potential ALCD-based therapeutic approaches.

Alteration of a specific microRNA regulating neprilysin expression in Alzheimer's disease.

Alzheimer's disease (AD) is characterized by accumulation of amyloid β (Aβ) plaques and aggregates of hyperphosphorylated tau protein. Genetic, epigenetic, and environmental factors play critical roles in AD etiology (Maloney & Lahiri-Lancet Neurol-15:760-774); however, underlying mechanisms of pathogenesis remain unclear. Both Aβ biogenesis pathways, involving APP and BACE1, and degradation pathways, involving membrane metallo-endopeptidase (MME), aka neprilysin, play critical roles in Aβ accumulation leading to AD. We posit that AD results from dysregulation of key biochemical pathways, particularly those involving non-coding microRNA (miRNA) (Chopra et al, Mol. Psychiatry-2020). We hypothesize that since MME is an important Aβ-clearing enzyme, disruption of miRNA regulation of MME would contribute to AD. Bioinformatics prediction revealed miRNA binding sites on the MME mRNA 3'-untranslated region (3'-UTR). Neuronally differentiated human neuroblastoma (SK-N-SH) cells were transfected with either siRNA against MME, candidate miRNA mimics, miRNA-inhibitors or negative controls. Levels of MME mRNA and protein were measured by qRT-PCR and western blotting, respectively. miR-181d emerged as a functional candidate. We tested sequence specificity and miR-181d activity with reporter gene- MME 3'-UTR fusion constructs. Levels of miR-181d were quantified in human brain tissue specimens from control, MCI, and AD subjects. Several human-specific miRNAs were predicted to target the MME 3'-UTR, including miR-9, miR-181d and miR-216. Notably, miR-181d treatment produced a significant reduction of MME mRNA and protein levels in neuronal cultures. Functional assays demonstrated miR-181d activity. MME is expressed in several human organs, the highest in kidney. Levels of miR-181d significantly influenced probability of AD diagnosis in human brain samples from temporal lobe and cerebellum but not posterior cingulate cortex. Overall, as miR-181d levels increased, likelihood of AD increased. This trend was particularly strong in female subjects vs. male subjects. Presence of the APOEε4 allele also increased likelihood of AD diagnosis, independent of miR-181d levels. In human neuronal cultures, miR-181d can down-regulate MME mRNA and protein, potentially through the MME 3'-UTR. Also, miR-181d is elevated in AD brains in multiple regions. Since MME is a key Aβ-degrading enzyme, its dysregulation by (presumably excess) miR-181d could contribute to AD pathogenesis. Supported by the NIA/NIH grant P01AG014449 to DKL and SEC.

Local thermal environment and warming influence supercooling and drive widespread shifts in the metabolome of diapausing Pieris rapae butterflies.

Global climate change has the potential to negatively impact biological systems as organisms are exposed to novel temperature regimes. Increases in annual mean temperature have been accompanied by disproportionate rates of change in temperature across seasons, and winter is the season warming most rapidly. Yet, we know relatively little about how warming will alter the physiology of overwintering organisms. Here, we simulated future warming conditions by comparing diapausing Pieris rapae butterfly pupae collected from disparate thermal environments and by exposing P. rapae pupae to acute and chronic increases in temperature. First, we compared internal freezing temperatures (supercooling points) of diapausing pupae that were developed in common-garden conditions but whose parents were collected from northern Vermont, USA, or North Carolina, USA. Matching the warmer winter climate of North Carolina, North Carolina pupae had significantly higher supercooling points than Vermont pupae. Next, we measured the effects of acute and chronic warming exposure in Vermont pupae and found that warming induced higher supercooling points. We further characterized the effects of chronic warming by profiling the metabolomes of Vermont pupae via untargeted LC-MS metabolomics. Warming caused significant changes in abundance of hundreds of metabolites across the metabolome. Notably, there were warming-induced shifts in key biochemical pathways, such as pyruvate metabolism, fructose and mannose metabolism, and β-alanine metabolism, suggesting shifts in energy metabolism and cryoprotection. These results suggest that warming affects various aspects of overwintering physiology in P. rapae and may be detrimental depending on the frequency and variation of winter warming events. Further research is needed to ascertain the extent to which the effects of warming are felt among a broader set of populations of P. rapae, and among other species, in order to better predict how insects may respond to changes in winter thermal environments.

MicroRNA Targeting.

MicroRNAs (miRNAs) are small noncoding elements that play essential roles in the posttranscriptional regulation of biochemical processes. miRNAs recognize and target multiple mRNAs; therefore, investigating miRNA dysregulation is an indispensable strategy to understand pathological conditions and to design innovative drugs. Targeting miRNAs in diseases improve outcomes of several therapeutic strategies thus, this present study highlights miRNA targeting methods through experimental assays and bioinformatics tools. The first part of this review focuses on experimental miRNA targeting approaches for elucidating key biochemical pathways. A growing body of evidence about the miRNA world reveals the fact that it is not possible to uncover these molecules' structural and functional characteristics related to the biological processes with a deterministic approach. Instead, a systemic point of view is needed to truly understand the facts behind the natural complexity of interactions and regulations that miRNA regulations present. This task heavily depends both on computational and experimental capabilities. Fortunately, several miRNA bioinformatics tools catering to nonexperts are available as complementary wet-lab approaches. For this purpose, this work provides recent research and information about computational tools for miRNA targeting research.

Effects of Organic Fertilizers on the Soil Microorganisms Responsible for N2O Emissions: A Review.

The use of organic fertilizers constitutes a sustainable strategy to recycle nutrients, increase soil carbon (C) stocks and mitigate climate change. Yet, this depends largely on balance between soil C sequestration and the emissions of the potent greenhouse gas nitrous oxide (N2O). Organic fertilizers strongly influence the microbial processes leading to the release of N2O. The magnitude and pattern of N2O emissions are different from the emissions observed from inorganic fertilizers and difficult to predict, which hinders developing best management practices specific to organic fertilizers. Currently, we lack a comprehensive evaluation of the effects of OFs on the function and structure of the N cycling microbial communities. Focusing on animal manures, here we provide an overview of the effects of these organic fertilizers on the community structure and function of nitrifying and denitrifying microorganisms in upland soils. Unprocessed manure with high moisture, high available nitrogen (N) and C content can shift the structure of the microbial community, increasing the abundance and activity of nitrifying and denitrifying microorganisms. Processed manure, such as digestate, compost, vermicompost and biochar, can also stimulate nitrifying and denitrifying microorganisms, although the effects on the soil microbial community structure are different, and N2O emissions are comparatively lower than raw manure. We propose a framework of best management practices to minimize the negative environmental impacts of organic fertilizers and maximize their benefits in improving soil health and sustaining food production systems. Long-term application of composted manure and the buildup of soil C stocks may contribute to N retention as microbial or stabilized organic N in the soil while increasing the abundance of denitrifying microorganisms and thus reduce the emissions of N2O by favoring the completion of denitrification to produce dinitrogen gas. Future research using multi-omics approaches can be used to establish key biochemical pathways and microbial taxa responsible for N2O production under organic fertilization.

Role of microRNAs in Pressure Ulcer Immune Response, Pathogenesis, and Treatment.

Pressure ulcers are preventable, yet highly prevalent, chronic wounds that have significant patient morbidity and high healthcare costs. Like other chronic wounds, they are characterized by impaired wound healing due to dysregulated immune processes. This review will highlight key biochemical pathways in the pathogenesis of pressure injury and how this signaling leads to impaired wound healing. This review is the first to comprehensively describe the current literature on microRNA (miRNA, miR) regulation of pressure ulcer pathophysiology.

Targeted toxicometabolomics of endosulfan sulfate in adult zebrafish (Danio rerio) using GC-MS/MS in multiple reaction monitoring mode

Endosulfan sulfate is a major oxidative metabolite of the chlorinated insecticide endosulfan. In this study, a targeted metabolomics approach was used to investigate the toxic mechanisms of endosulfan sulfate in adult zebrafish using the multiple reaction monitoring mode of a GC-MS/MS. The LC50 of endosulfan sulfate in adult zebrafish was determined and then zebrafish were exposed to endosulfan sulfate at one-tenth the LC50 (0.1LC50) or the LC50 for 24 and 48 h. After exposure, the fish were extracted, derivatized and analyzed by GC-MS/MS for 379 metabolites to identify 170 metabolites. Three experimental groups (control, 0.1LC50 and LC50) were clearly separated in PLS-DA score plots. Based on the VIP, ANOVA, and fold change results, 40 metabolites were selected as biomarkers. Metabolic pathways associated with those metabolites were identified using MetaboAnalyst 4.0 as follows: aminoacyl-tRNA biosynthesis, valine/leucine/isoleucine biosynthesis, citrate cycle, glycerolipid metabolism, and arginine/proline metabolism. Gene expression studies confirmed the activation of citrate cycle and glycerolipids metabolism. MDA levels of the exposed group significantly increased in oxidative toxicity assay tests. Such significant perturbations of important metabolites within key biochemical pathways must result in biologically hazardous effects in zebrafish.