Biomarker identification and risk prediction model development for differentiated thyroid carcinoma lung metastasis based on primary lesion proteomics.

PURPOSE: The rising global high incidence of differentiated thyroid carcinoma (DTC) has led to a significant increase in patients presenting with lung metastasis of DTC (LMDTC). This population poses a significant challenge in clinical practice, necessitating the urgent development of effective risk stratification methods and predictive tools for lung metastasis. EXPERIMENTAL DESIGN: Through proteomic analysis of large samples of primary lesion and dual validation employing parallel reaction monitoring and immunohistochemistry, we identified eight hub proteins as potential biomarkers. By expanding the sample size and conducting statistical analysis on clinical features and hub protein expression, we constructed three risk prediction models. RESULTS: This study identified eight hub proteins-SUCLG1/2, DLAT, IDH3B, ACSF2, ACO2, CYCS and VDAC2- as potential biomarkers for predicting DTC lung metastasis risk. We developed and internally validated three risk prediction models incorporating both clinical characteristics and hub protein expression. Our findings demonstrated that the combined prediction model exhibited optimal predictive performance, with the highest discrimination (AUC: 0.986) and calibration (Brier score: 0.043). Application of the combined prediction model within a specific risk threshold (0-0.97) yielded maximal clinical benefit. Finally, we constructed a nomogram based on the combined prediction model. CONCLUSIONS: As a large sample size study in lung metastatic DTC research, the identification of biomarkers through primary lesion proteomics and the development of risk prediction models integrating clinical features and hub protein biomarkers offer valuable insights for predicting DTC lung metastasis and establishing personalised treatment strategies.

Rapid molecular identification of Rana dybowskii by species-specific primers.

Oviductus Ranae is the dried oviduct from Rana dybowskii, a forest frog species with medicinal, tonic, and cosmetic properties. Due to the high price and resource shortage, counterfeit varieties of Oviductus Ranae often appear in the market. However, traditional identification methods cannot accurately differentiate between Oviductus Ranae and its adulterants. In this study, a rapid molecular identification method has been established. The method involves extracting genomic DNA in just 30 s using filter paper purification, species-specific rapid polymerase chain reaction (PCR) amplification, and finally, fluorescence detection of the products. It can accurately identify Oviductus Ranae and its three common adulterants in about 30 min, making the process simple, fast, and highly specific.

Microglial activation in the lateral amygdala promotes anxiety-like behaviors in mice with chronic moderate noise exposure.

BACKGROUND: Long-term non-traumatic noise exposure, such as heavy traffic noise, can elicit emotional disorders in humans. However, the underlying neural substrate is still poorly understood. METHODS: We exposed mice to moderate white noise for 28 days to induce anxiety-like behaviors, measured by open-field, elevated plus maze, and light-dark box tests. In vivo multi-electrode recordings in awake mice were used to examine neuronal activity. Chemogenetics were used to silence specific brain regions. Viral tracing, immunofluorescence, and confocal imaging were applied to define the neural circuit and characterize the morphology of microglia. RESULTS: Exposure to moderate noise for 28 days at an 85-dB sound pressure level resulted in anxiety-like behaviors in open-field, elevated plus maze, and light-dark box tests. Viral tracing revealed that fibers projecting from the auditory cortex and auditory thalamus terminate in the lateral amygdala (LA). A noise-induced increase in spontaneous firing rates of the LA and blockade of noise-evoked anxiety-like behaviors by chemogenetic inhibition of LA glutamatergic neurons together confirmed that the LA plays a critical role in noise-induced anxiety. Noise-exposed animals were more vulnerable to anxiety induced by acute noise stressors than control mice. In addition to these behavioral abnormalities, ionized calcium-binding adaptor molecule 1 (Iba-1)-positive microglia in the LA underwent corresponding morphological modifications, including reduced process length and branching and increased soma size following noise exposure. Treatment with minocycline to suppress microglia inhibited noise-associated changes in microglial morphology, neuronal electrophysiological activity, and behavioral changes. Furthermore, microglia-mediated synaptic phagocytosis favored inhibitory synapses, which can cause an imbalance between excitation and inhibition, leading to anxiety-like behaviors. CONCLUSIONS: Our study identifies LA microglial activation as a critical mediator of noise-induced anxiety-like behaviors, leading to neuronal and behavioral changes through selective synapse phagocytosis. Our results highlight the pivotal but previously unrecognized roles of LA microglia in chronic moderate noise-induced behavioral changes.

Dissociable contributions of the hippocampus and orbitofrontal cortex to representing task space in a social context.

The hippocampus (HC) and the orbitofrontal cortex (OFC) jointly encode a map-like representation of a task space to guide behavior. It remains unclear how the OFC and HC interact in encoding this map-like representation, though previous studies indicated that both regions have different functions. We acquired the functional magnetic resonance imaging data under a social navigation task in which participants interacted with characters in a two-dimensional "social space." We calculate the social relationships between the participants and characters and used a drift-diffusion model to capture the inner process of social interaction. Then we used multivoxel pattern analysis to explore the brain-behavior relationship. We found that (i) both the HC and the OFC showed higher activations during the selective trial than the narrative trial; (ii) the neural pattern of the right HC was associated with evidence accumulation during social interaction, and the pattern of the right lateral OFC was associated with the social relationship; (iii) the neural pattern of the HC can decode the participants choices, while the neural pattern of the OFC can decode the task information about trials. The study provided evidence for distinct roles of the HC and the OFC in encoding different information when representing social space.

Characterization of Anxiety-Like Behaviors and Neural Circuitry following Chronic Moderate Noise Exposure in Mice.

BACKGROUND: Commonly encountered nontraumatic, moderate noise is increasingly implicated in anxiety; however, the neural substrates underlying this process remain unclear. OBJECTIVES: We investigated the neural circuit mechanism through which chronic exposure to moderate-level noise causes anxiety-like behaviors. METHODS: Mice were exposed to chronic, moderate white noise [85 decibel (dB) sound pressure level (SPL)], 4 h/d for 4 wk to induce anxiety-like behaviors, which were assessed by open field, elevated plus maze, light-dark box, and social interaction tests. Viral tracing, immunofluorescence confocal imaging, and brain slice patch-clamp recordings were used to characterize projections from auditory brain regions to the lateral amygdala. Neuronal activities were characterized by in vivo multielectrode and fiber photometry recordings in awake mice. Optogenetics and chemogenetics were used to manipulate specific neural circuitry. RESULTS: Mice chronically (4 wk) exposed to moderate noise (85 dB SPL, 4 h/d) demonstrated greater neuronal activity in the lateral amygdala (LA), and the LA played a critical role in noise-induced anxiety-like behavior in these model mice. Viral tracing showed that the LA received monosynaptic projections from the medial geniculate body (MG) and auditory cortex (ACx). Optogenetic excitation of the MG→LA or ACx→LA circuits acutely evoked anxiety-like behaviors, whereas their chemogenetic inactivation abolished noise-induced anxiety-like behavior. Moreover, mice chronically exposed to moderate noise were more susceptible to acute stress, with more neuronal firing in the LA, even after noise withdrawal. DISCUSSION: Mice exposed to 4 wk of moderate noise (85 dB SPL, 4 h/d) demonstrated behavioral and physiological differences compared to controls. The neural circuit mechanisms involved greater excitation from glutamatergic neurons of the MG and ACx to LA neurons under chronic, moderate noise exposure, which ultimately promoted anxiety-like behaviors. Our findings support the hypothesis that nontraumatic noise pollution is a potentially serious but unrecognized public health concern. https://doi.org/10.1289/EHP12532.

A neural circuit for gastric motility disorders driven by gastric dilation in mice.

Introduction: Symptoms of gastric motility disorders are common clinical manifestations of functional gastrointestinal disorders (FGIDs), and are triggered and exacerbated by stress, but the neural pathways underpinning them remain unclear. Methods: We set-up a mouse model by gastric dilation (GD) in which the gastric dynamics were assessed by installing strain gauges on the surface of the stomach. The neural pathway associated with gastric motility disorders was investigated by behavioral tests, electrophysiology, neural circuit tracing, and optogenetics and chemogenetics involving projections of the corticotropin-releasing hormone (CRH) from the paraventricular nucleus of the hypothalamus (PVN) to acetylcholine (ChAT) neurons in the dorsal motor nucleus of the vagus (DMV). Results: We found that GD induced gastric motility disorders were accompanied by activation of PVN CRH neurons, which could be alleviated by strategies that inhibits the activity of PVN CRH neurons. In addition, we identified a neural pathway in which PVN CRH neurons project into DMV ChAT neurons, modulated activity of the PVN CRH →DMV ChAT pathway to alleviate gastric motility disorders induced by GD. Discussion: These findings indicate that the PVN CRH →DMV ChAT pathway may mediate at least some aspects of GD related gastric motility, and provide new insights into the mechanisms by which somatic stimulation modulates the physiological functions of internal organs and systems.

Green light induces antinociception via visual-somatosensory circuits.

Light has been shown to relieve pain, but the underlying neural mechanisms remain unknown. Here, we show that low-intensity (200 lux) green light treatment exerts antinociceptive effects through a neural circuit from the visual cortex projecting to the anterior cingulate cortex (ACC) in mice. Specifically, viral tracing, in vivo two-photon calcium imaging, and fiber photometry recordings show that green light activated glutamatergic projections from the medial part of the secondary visual cortex (V2MGlu) to GABAergic neurons in the ACC, which drives inhibition of local glutamatergic neurons (V2MGlu→ACCGABA→Glu). Optogenetic or chemogenetic activation of the V2MGlu→ACCGABA→Glu circuit mimics green-light-induced antinociception in both neuropathic and inflammatory pain model mice. Artificial inhibition of ACC-projecting V2MGlu neurons abolishes the antinociception induced by green light. Taken together, our study shows the V2M-ACC circuit as a potential candidate mediating green-light-induced antinociceptive effects.

Endogenous hydrogen sulfide promotes human preimplantation embryonic development by regulating metabolism-related gene expression.

Hydrogen sulfide (H2S) as an endogenous gaseous signaling molecule had been proved to play a vital role in gametes physiology, covering meiosis, maturation and aging. However, little is known about H2S involvement in embryonic development. The present study explored the positive effect of H2S on human early embryonic development. Results validated that the two H2S producing enzymes, CBS and CSE mRNA and proteins were identified in donated human cleavage and blastocyst-stage embryos. The l-cysteine incubation produced endogenous H2S in human blastocysts. NaHS positively affected in vitro blastulation. Single-cell RNA-seq analysis identified 228 differentially expressed genes (DEGs) after NaHS treatment versus the control. The Gene Ontology (GO) enrichment analysis of DEGs showed that genes for protein modification and metabolism were significantly enriched in the NaHS treatment group. For the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, 2-oxocarboxylic acid metabolism, glycosaminoglycan biosynthesis-keratan sulfate, steroid biosynthesis, carbon metabolism, and biosynthesis of amino acids were significantly enriched. Six DEGs, including Neural EGFL like 1 (NELL1), aconitase 1 (ACO1), phosphoglycerate mutase 1 (PGAM1), TP53 induced glycolysis regulatory phosphatase (TIGAR), UDP-GlcNAc:betaGal beta-1,3-N-acetylglucosaminyltransferase 2 (B3GNT2), and carbohydrate Sulfotransferase 4 (CHST4) were validate by real-time RT-PCR. These findings suggest that H2S is a positive regulator of early embryonic development and may alter the transcription of embryonic genes for protein modification and metabolism in human embryos.

Early-life inflammation promotes depressive symptoms in adolescence via microglial engulfment of dendritic spines.

Early-life inflammation increases the risk for depression in later life. Here, we demonstrate how early-life inflammation causes adolescent depressive-like symptoms: by altering the long-term neuronal spine engulfment capacity of microglia. For mice exposed to lipopolysaccharide (LPS)-induced inflammation via the Toll-like receptor 4/NF-κB signaling pathway at postnatal day (P) 14, ongoing longitudinal imaging of the living brain revealed that later stress (delivered during adolescence on P45) increases the extent of microglial engulfment around anterior cingulate cortex (ACC) glutamatergic neuronal (ACCGlu) spines. When the ACC microglia of LPS-treated mice were deleted or chemically inhibited, the mice did not exhibit depressive-like behaviors during adolescence. Moreover, we show that the fractalkine receptor CX3CR1 mediates stress-induced engulfment of ACCGlu neuronal spines. Together, our findings establish that early-life inflammation causes dysregulation of microglial engulfment capacity, which encodes long-lasting maladaptation of ACCGlu neurons to stress, thus promoting development of depression-like symptoms during adolescence.

Synthesis and Mechanism of High Structural Stability of Nickel-Rich Cathode Materials by Adjusting Li-Excess.

It has been a long-term challenge to improve the phase stability of Ni-rich LiNixMnyCo1-x-yO2 (x ≥ 0.6) transition metal (TM) oxides for large-scale applications. Herein, a new structure engineering strategy is utilized to optimize the structural arrangement of Li1+x(Ni0.88Mn0.06Co0.06)1-xO2 (NMC88) with a different Li-excess content. It was found that structure stability and particle sizes can be tuned with suitable Li-excess contents. NMC88 with an actual Li-excess of 2.7% (x = 0.027, Li/TM = 1.055) exhibits a high discharge capacity (209.1 mAh g-1 at 3.0-4.3 V, 0.1 C) and maintains 91.7% after the 100th cycle at 1 C compared with the NMC88 sample free of Li-excess. It also performs a delayed voltage decay and a good rate capacity, delivering 145.8 mAh g-1 at a high rate of 10 C. Multiscale characterization technologies including ex/in situ X-ray diffraction (XRD), focused ion beam (FIB) cutting-scanning electronic microscopy (SEM), and transmission electron microscopy (TEM) results show that a proper Li-excess (2.7%) content contributes to the formation of a broader Li slab, optimized cation mixing ratio, and even particle sizes. Therefore, NMC88 with a proper Li-excess is a good choice for next-generation cathode materials.

Ultrastructural observation of mesophyll cells and temporal expression profiles of the genes involved in transitory starch metabolism in flag leaves of wheat after anthesis.

Transitory starch in cereal plant leaves is synthesized during the day and remobilized at night to provide a carbon source for growth and grain filling, but its mechanistic basis is still poorly understood. The objective of this study is to explore the regulatory mechanism for starch biosynthesis and degradation in plant source organs. Using transmission electron microscopy, we observed that during the day after anthesis, starch granules in mesophyll cells of wheat flag leaves accumulated in chloroplasts and the number of starch granules gradually decreased with wheat leaf growth. During the night, starch granules synthesized in chloroplasts during the day were completely or partially degraded. The transcript levels of 26 starch synthesis-related genes and 16 starch breakdown-related genes were further measured using quantitative real-time reverse transcription polymerase chain reaction. Expression profile analysis revealed that starch metabolism genes were clustered into two groups based on their temporal expression patterns. The genes in the first group were highly expressed and presumed to play crucial roles in starch metabolism. The genes in the other group were not highly expressed in flag leaves and may have minor functions in starch metabolism in leaf tissue. The functions of most of these genes in leaves were further discussed. The starch metabolism-related genes that are predominantly expressed in wheat flag leaves differ from those expressed in wheat grain, indicating that two different pathways for starch metabolism operate in these tissues. This provides specific information on the molecular mechanisms of transitory starch metabolism in higher plants.

Increasing the starch content and grain weight of common wheat by overexpression of the cytosolic AGPase large subunit gene.

ADP-glucose pyrophosphorylase (AGPase) catalyzes the first committed step of starch synthesis. AGPase is a heterotetramer composed of two large subunits and two small subunits, has cytosolic and plastidial isoforms, and is detected mainly in the cytosol of endosperm in cereal crops. To investigate the effects of AGPase cytosolic large subunit gene (LSU I) on starch biosynthesis in higher plant, in this study, a TaLSU I gene from wheat was overexpressed under the control of an endosperm-specific promoter in a wheat cultivar (Yumai 34). PCR, Southern blot, and real-time RT-PCR analyses indicated that the transgene was integrated into the genome of transgenic plants and was overexpressed in their progeny. The overexpression of the TaLSU I gene remarkably enhanced AGPase activity, endosperm starch weight, grain number per spike, and single grain weight. Therefore, we conclude that overexpression of the TaLSU I gene enhances the starch biosynthesis in endosperm of wheat grains, having potential applications in wheat breeding to develop a high-yield wheat cultivar with high starch weight and kernel weight.

Proteomic analysis of leaves and roots of common wheat (Triticum aestivum L.) under copper-stress conditions.

Proteomic studies were performed to identify the protein species involved in copper (Cu) stress responses in common wheat. Two-week-old wheat seedlings were exposed to 100 µM CuSO4 treatment for 3 days. Growth of shoots and roots was markedly inhibited and lipid peroxidation was greatly increased. Cu was readily absorbed by wheat seedlings, with greater Cu contents in roots than in leaves. Using 2-DE method, 98 protein spots showed significantly enhanced or reduced abundance, of which 93 were successfully identified. Of these identified protein species, 49 and 44 were found in roots and leaves, respectively. Abundance of most of identified protein species, which function in signal transduction, stress defense, and energy production, was significantly enhanced, while that of many protein species involved in carbohydrate metabolism, protein metabolism, and photosynthesis was severely reduced. The Cu-responsive protein interaction network revealed 36 key proteins, most of which may be regulated by abscisic acid (ABA), ethylene, jasmonic acid (JA), and so on. Exogenous JA application showed a protective effect against Cu stress and significantly increased transcripts of the glutathione S-transferase (GST) gene. This study provides insight into the molecular mechanisms of Cu responses in higher plants.

Salicylic acid increases the contents of glutathione and ascorbate and temporally regulates the related gene expression in salt-stressed wheat seedlings.

Exogenous salicylic acid (SA) significantly improved abiotic tolerance in higher plants, and ascorbate (ASA) and glutathione (GSH) play important roles in abiotic tolerance. In this study, SA (0.5mM) markedly increased the contents of ASA and GSH in SA-treated plants during salt stress (250mM NaCl). The transcript levels of the genes encoding ASA and GSH cycle enzymes were measured using quantitative real-time PCR. The results indicated that, during salt stress, exogenous SA significantly enhanced the transcripts of glutathione peroxidase (GPX1), phospholipid hydroperoxide glutathione peroxidase (GPX2) and dehydroascorbate reductase (DHAR) genes at 12h, glutathione reductase (GR) at 24h, 48h and 72h, glutathione-S-transferase 1 (GST1), 2 (GST2), monodehydroascorbate reductase (MDHAR) and glutathione synthetase (GS) at the 48h and 72h after salt stress, respectively. The results implied that SA temporally regulated the transcript levels of the genes encoding ASA-GSH cycle enzymes, resulting in the increased contents of GSH and ASA and enhanced salt tolerance.

Comprehensive analysis of the transcription of starch synthesis genes and the transcription factor RSR1 in wheat (Triticum aestivum) endosperm.

The cDNA sequences of 26 starch synthesis genes were identified in common wheat (Triticum aestivum L.), and their transcript levels were measured using quantitative real-time RT-PCR to assess the function of individual genes and the regulatory mechanism in wheat endosperm. The expression patterns of 26 genes in wheat endosperm were classified into three groups. The genes in group 1 were richly expressed in the early stage of grain development and may be involved in the construction of fundamental cell machinery, synthesis of glucan primers, and initiation of starch granules. The genes in group 2 were highly expressed during the middle and late stages of grain development, and their expression profiles were similar to the accumulation rate of endosperm starch; these genes are presumed to play a crucial role in starch production. The genes in group 3 were scantily expressed throughout the grain development period and might be associated with transitory starch synthesis. Transcripts of the negative transcription factor TaRSR1 were high at the early and late stages of grain development but low during the middle stage. The expression pattern of TaRSR1 was almost opposite to those of the group 2 starch synthesis genes, indicating that TaRSR1 might negatively regulate the expression of many endosperm starch synthesis genes during grain development.

Proteomics reveals the effects of salicylic acid on growth and tolerance to subsequent drought stress in wheat.

Pretreatment with 0.5 mM salicylic acid (SA) for 3 days significantly enhanced the growth and tolerance to subsequent drought stress (PEG-6000, 15%) in wheat seedlings, manifesting as increased shoot and root dry weights, and decreased lipid peroxidation. Total proteins from wheat leaves exposed to (i) 0.5 mM SA pretreatment, (ii) drought stress, and (iii) 0.5 mM SA treatment plus drought-stress treatments were analyzed using a proteomics method. Eighty-two stress-responsive protein spots showed significant changes, of which 76 were successfully identified by MALDI-TOF-TOF. Analysis of protein expression patterns revealed that proteins associated with signal transduction, stress defense, photosynthesis, carbohydrate metabolism, protein metabolism, and energy production could by involved in SA-induced growth and drought tolerance in wheat seedlings. Furthermore, the SA-responsive protein interaction network revealed 35 key proteins, suggesting that these proteins are critical for SA-induced tolerance.