Genomics-based analysis of four porcine-derived lactic acid bacteria strains and their evaluation as potential probiotics.

The search for probiotics and exploration of their functions are crucial for livestock farming. Recently, porcine-derived lactic acid bacteria (LAB) have shown great potential as probiotics. However, research on the evaluation of porcine-derived LAB as potential probiotics through genomics-based analysis is relatively limited. The present study analyzed four porcine-derived LAB strains (Lactobacillus johnsonii L16, Latilactobacillus curvatus ZHA1, Ligilactobacillus salivarius ZSA5 and Ligilactobacillus animalis ZSB1) using genomic techniques and combined with in vitro tests to evaluate their potential as probiotics. The genome sizes of the four strains ranged from 1,897,301 bp to 2,318,470 bp with the GC contents from 33.03 to 41.97%. Pan-genomic analysis and collinearity analysis indicated differences among the genomes of four strains. Carbohydrate active enzymes analysis revealed that L. johnsonii L16 encoded more carbohydrate active enzymes than other strains. KEGG pathway analysis and in vitro tests confirmed that L. johnsonii L16 could utilize a wide range of carbohydrates and had good utilization capacity for each carbohydrate. The four strains had genes related to acid tolerance and were tolerant to low pH, with L. johnsonii L16 showing the greatest tolerance. The four strains contained genes related to bile salt tolerance and were able to tolerate 0.1% bile salt. Four strains had antioxidant related genes and exhibited antioxidant activity in in vitro tests. They contained the genes linked with organic acid biosynthesis and exhibited antibacterial activity against enterotoxigenic Escherichia coli K88 (ETEC K88) and Salmonella 6,7:c:1,5, wherein, L. johnsonii L16 and L. salivarius ZSA5 had gene clusters encoding bacteriocin. Results suggest that genome analysis combined with in vitro tests is an effective approach for evaluating different strains as probiotics. The findings of this study indicate that L. johnsonii L16 has the potential as a probiotic strain among the four strains and provide theoretical basis for the development of probiotics in swine production.

Thermally Stable Polymer-Rich Solid Electrolyte Interphase for Safe Lithium Metal Pouch Cells.

Serious safety risks caused by the high reactivity of lithium metal against electrolytes severely hamper the practicability of lithium metal batteries. By introducing unique polymerization site and more fluoride substitution, we built an in situ formed polymer-rich solid electrolyte interphase upon lithium anode to improve battery safety. The fluorine-rich and hydrogen-free polymer exhibits high thermal stability, which effectively reduces the continuous exothermic reaction between electrolyte and anode/cathode. As a result, the critical temperature for thermal safety of 1.0 Ah lithium-LiNi0.5 Co0.2 Mn0.3 O2 pouch cell can be increased from 143.2 °C to 174.2 °C. The more dangerous "ignition" point of lithium metal batteries, the starting temperature of battery thermal runaway, has been dramatically raised from 240.0 °C to 338.0 °C. This work affords novel strategies upon electrolyte design, aiming to pave the way for high-energy-density and thermally safe lithium metal batteries.

Behavioural tactic predicts preoptic-hypothalamic gene expression more strongly than developmental morph in fish with alternative reproductive tactics.

Reproductive success relies on the coordination of social behaviours, such as territory defence, courtship and mating. Species with extreme variation in reproductive tactics are useful models for identifying the neural mechanisms underlying social behaviour plasticity. The plainfin midshipman (Porichthys notatus) is a teleost fish with two male reproductive morphs that follow widely divergent developmental trajectories and display alternative reproductive tactics (ARTs). Type I males defend territories, court females and provide paternal care, but will resort to cuckoldry if they cannot maintain a territory. Type II males reproduce only through cuckoldry. We sought to disentangle gene expression patterns underlying behavioural tactic, in this case ARTs, from those solely reflective of developmental morph. Using RNA-sequencing, we investigated differential transcript expression in the preoptic area-anterior hypothalamus (POA-AH) of courting type I males, cuckolding type I males and cuckolding type II males. Unexpectedly, POA-AH differential expression was more strongly coupled to behavioural tactic than morph. This included a suite of transcripts implicated in hormonal regulation of vertebrate social behaviour. Our results reveal that divergent expression patterns in a conserved neuroendocrine centre known to regulate social-reproductive behaviours across vertebrate lineages may be uncoupled from developmental history to enable plasticity in the performance of reproductive tactics.

Gene expression underlying enhanced, steroid-dependent auditory sensitivity of hair cell epithelium in a vocal fish.

BACKGROUND: Successful animal communication depends on a receiver's ability to detect a sender's signal. Exemplars of adaptive sender-receiver coupling include acoustic communication, often important in the context of seasonal reproduction. During the reproductive summer season, both male and female midshipman fish (Porichthys notatus) exhibit similar increases in the steroid-dependent frequency sensitivity of the saccule, the main auditory division of the inner ear. This form of auditory plasticity enhances detection of the higher frequency components of the multi-harmonic, long-duration advertisement calls produced repetitively by males during summer nights of peak vocal and spawning activity. The molecular basis of this seasonal auditory plasticity has not been fully resolved. Here, we utilize an unbiased transcriptomic RNA sequencing approach to identify differentially expressed transcripts within the saccule's hair cell epithelium of reproductive summer and non-reproductive winter fish. RESULTS: We assembled 74,027 unique transcripts from our saccular epithelial sequence reads. Of these, 6.4 % and 3.0 % were upregulated in the reproductive and non-reproductive saccular epithelium, respectively. Gene ontology (GO) term enrichment analyses of the differentially expressed transcripts showed that the reproductive saccular epithelium was transcriptionally, translationally, and metabolically more active than the non-reproductive epithelium. Furthermore, the expression of a specific suite of candidate genes, including ion channels and components of steroid-signaling pathways, was upregulated in the reproductive compared to the non-reproductive saccular epithelium. We found reported auditory functions for 14 candidate genes upregulated in the reproductive midshipman saccular epithelium, 8 of which are enriched in mouse hair cells, validating their hair cell-specific functions across vertebrates. CONCLUSIONS: We identified a suite of differentially expressed genes belonging to neurotransmission and steroid-signaling pathways, consistent with previous work showing the importance of these characters in regulating hair cell auditory sensitivity in midshipman fish and, more broadly, vertebrates. The results were also consistent with auditory hair cells being generally more physiologically active when animals are in a reproductive state, a time of enhanced sensory-motor coupling between the auditory periphery and the upper harmonics of vocalizations. Together with several new candidate genes, our results identify discrete patterns of gene expression linked to frequency- and steroid-dependent plasticity of hair cell auditory sensitivity.

Neural transcriptome reveals molecular mechanisms for temporal control of vocalization across multiple timescales.

BACKGROUND: Vocalization is a prominent social behavior among vertebrates, including in the midshipman fish, an established model for elucidating the neural basis of acoustic communication. Courtship vocalizations produced by territorial males are essential for reproductive success, vary over daily and seasonal cycles, and last up to hours per call. Vocalizations rely upon extreme synchrony and millisecond precision in the firing of a homogeneous population of motoneurons, the vocal motor nucleus (VMN). Although studies have identified neural mechanisms driving rapid, precise, and stable neuronal firing over long periods of calling, little is known about underlying genetic/molecular mechanisms. RESULTS: We used RNA sequencing-based transcriptome analyses to compare patterns of gene expression in VMN to the surrounding hindbrain across three daily and seasonal time points of high and low sound production to identify candidate genes that underlie VMN's intrinsic and network neuronal properties. Results from gene ontology enrichment, enzyme pathway mapping, and gene category-wide expression levels highlighted the importance of cellular respiration in VMN function, consistent with the high energetic demands of sustained vocal behavior. Functionally important candidate genes upregulated in the VMN, including at time points corresponding to high natural vocal activity, encode ion channels and neurotransmitter receptors, hormone receptors and biosynthetic enzymes, neuromodulators, aerobic respiration enzymes, and antioxidants. Quantitative PCR and RNA-seq expression levels for 28 genes were significantly correlated. Many candidate gene products regulate mechanisms of neuronal excitability, including those previously identified in VMN motoneurons, as well as novel ones that remain to be investigated. Supporting evidence from previous studies in midshipman strongly validate the value of transcriptomic analyses for linking genes to neural characters that drive behavior. CONCLUSIONS: Transcriptome analyses highlighted a suite of molecular mechanisms that regulate vocalization over behaviorally relevant timescales, spanning milliseconds to hours and seasons. To our knowledge, this is the first comprehensive characterization of gene expression in a dedicated vocal motor nucleus. Candidate genes identified here may belong to a conserved genetic toolkit for vocal motoneurons facing similar energetic and neurophysiological demands.

Loss of T cell progenitor checkpoint control underlies leukemia initiation in Rag1-deficient nonobese diabetic mice.

NOD mice exhibit major defects in the earliest stages of T cell development in the thymus. Genome-wide genetic and transcriptome analyses were used to investigate the origins and consequences of an early T cell developmental checkpoint breakthrough in Rag1-deficient NOD mice. Quantitative trait locus analysis mapped the presence of checkpoint breakthrough cells to several known NOD diabetes susceptibility regions, particularly insulin-dependent diabetes susceptibility genes (Idd)9/11 on chromosome 4, suggesting common genetic origins for T cell defects affecting this trait and autoimmunity. Genome-wide RNA deep-sequencing of NOD and B6 Rag1-deficient thymocytes revealed the effects of genetic background prior to breakthrough, as well as the cellular consequences of the breakthrough. Transcriptome comparison between the two strains showed enrichment in differentially expressed signal transduction genes, prominently tyrosine kinase and actin-binding genes, in accord with their divergent sensitivities to activating signals. Emerging NOD breakthrough cells aberrantly expressed both stem cell-associated proto-oncogenes, such as Lmo2, Hhex, Lyl1, and Kit, which are normally repressed at the commitment checkpoint, and post-ß-selection checkpoint genes, including Cd2 and Cd5. Coexpression of genes characteristic of multipotent progenitors and more mature T cells persists in the expanding population of thymocytes and in the thymic leukemias that emerge with age in these mice. These results show that Rag1-deficient NOD thymocytes have T cell defects that can collapse regulatory boundaries at two early T cell checkpoints, which may predispose them to both leukemia and autoimmunity.

Melatonin action in a midbrain vocal-acoustic network.

Melatonin is a well-documented time-keeping hormone that can entrain an individual's physiology and behavior to the day-night cycle, though surprisingly little is known about its influence on the neural basis of social behavior, including vocalization. Male midshipman fish (Porichthys notatus) produce several call types distinguishable by duration and by daily and seasonal cycles in their production. We investigated melatonin's influence on the known nocturnal- and breeding season-dependent increase in excitability of the midshipman's vocal network (VN) that directly patterns natural calls. VN output is readily recorded from the vocal nerve as a 'fictive call'. Five days of constant light significantly increased stimulus threshold levels for calls electrically evoked from vocally active sites in the medial midbrain, supporting previous findings that light suppresses VN excitability, while 2-iodomelatonin (2-IMel; a melatonin analog) implantation decreased threshold. 2-IMel also increased fictive call duration evoked from medial sites as well as lateral midbrain sites that produced several-fold longer calls irrespective of photoregime or drug treatment. When stimulus intensity was incrementally increased, 2-IMel increased duration only at lateral sites, suggesting that melatonin action is stronger in the lateral midbrain. For animals receiving 5 days of constant darkness, known to increase VN excitability, systemic injections of either of two mammalian melatonin receptor antagonists increased threshold and decreased duration for calls evoked from medial sites. Our results demonstrate melatonin modulation of VN excitability and suggest that social context-dependent call types differing in duration may be determined by neuro-hormonal action within specific regions of a midbrain vocal-acoustic network.

Research on the characteristics and influencing factors of the spatial correlation network of cultivated land utilization ecological efficiency in the upper reaches of the Yangtze River, China.

Researching the structural characteristics of the spatial correlation network of cultivated land utilization ecological efficiency is of great significance to China's food security and agricultural green and low-carbon development. Taking 47 cities (autonomous prefectures) in the upper reaches of the Yangtze River as the research object, the ecological efficiency of cultivated land utilization from 2010 to 2020 was measured based on the unexpected output model (Super SBM), and the spatial correlation matrix was constructed using the revised gravity model. The structural characteristics of the spatial correlation network were analyzed using the social network model (SNA), and finally, the factors affecting the spatial correlation network of cultivated land utilization ecological efficiency in the upper reaches of the Yangtze River were analyzed through the quadratic assignment procedure (QAP) model. The results show that: (1) the ecological efficiency of cultivated land utilization in the upper reaches of the Yangtze River has been increasing year by year, but the overall level is low, and there is a large gap among provinces. Sichuan Province has the highest average value of 0.605, and Yunnan Province has the lowest average value of 0.359. (2) The ecological efficiency of cultivated land utilization in the upper reaches of the Yangtze River has broken through the provincial boundaries and has formed an obvious spatial correlation network, but the overall density is low, and the network is still relatively loose, needing further development and improvement. Chengdu, Yibin, Luzhou, and other cities are located in the center of the network and have formed four cohesive subgroups. (3)The differences in the level of agricultural economic development, the rural per capita disposable income, the differences in agricultural mechanization intensity, the regional population differences, and spatial adjacency have an impact on the spatial network of ecological efficiency of cultivated land utilization in the upper reaches of the Yangtze River. The difference in the level of agricultural economic development, the rural per capita disposable income, and the differences in agricultural mechanization intensity are negatively correlated, while the regional population differences are positively correlated with spatial adjacency.

The Fecal Redox Potential in Healthy and Diarrheal Pigs and Their Correlation with Microbiota.

The redox potential plays a critical role in sustaining the stability of gut microbiota. This study measured the fecal redox potential in healthy and diarrheal pigs using direct and dilution methods and investigated their correlation with microbiota. The results showed that the fluctuations in the redox potential of healthy pig feces were consistent using two different methods and the two methods are equivalent based on an equivalence test. The redox potential was positively correlated with the number of fungi and negatively related to the total bacteria. The relative or absolute abundances of many bacteria at the phyla and genus levels were associated with redox potential. In diarrheal pigs, the potentiometric trends of the two methods demonstrated an opposing pattern and the correlation with total bacteria was reversed. Precipitously elevated redox potential was detected post-diarrhea using dilution methods. The absolute abundance of Escherichia-Shigella and Fuurnierella was positively correlated with redox potential, while both relative and absolute abundances of Limosilactobacillus were positively correlated. These results suggest that both methods are suitable for detecting gut redox potential in healthy pigs, while the dilution method is more suitable for diarrheal pigs. The findings on the correlation of Limosilactobacillus, Prevotella, and Escherichia-Shigella with redox potential offer novel insights for targeted modulation of intestinal health.

Pectin supplementation accelerates post-antibiotic gut microbiome reconstitution orchestrated with reduced gut redox potential.

Antibiotic-induced gut dysbiosis (AID) presents a big challenge to host health, and the recovery from this dysbiosis is often slow and incomplete. AID is typically characterized by elevation in redox potential, Enterobacteriaceae load, and aerobic metabolism. In our previous study, a pectin-enriched diet was demonstrated to decrease fecal redox potential and modulate the gut microbiome. Therefore, we propose that pectin supplementation may modulate gut redox potential and favor post-antibiotic gut microbiome reconstitution from dysbiosis. In the present study, rats with AIDwere used to investigate the effects of pectin supplementation on post-antibiotic gut microbiome reconstitution from dysbiosis. The results showed that pectin supplementation accelerated post-antibiotic reconstitution of gut microbiome composition and function and led to enhancement of anabolic reductive metabolism and weakening of catabolic oxidative pathways. These results were corroborated by the measurement of redox potential, findings suggesting that pectin favors post-antibiotic recovery from dysbiosis. Pectin-modulated fecal microbiota transplantation accelerated the decrease in antibiotics-elevated redox potential and Enterobacteriaceae load similarly to pectin supplementation. Moreover, both pectin supplementation and Pectin-modulated fecal microbiota transplantation enriched anaerobic members, primarily from Lachnospiraceae orchestration with enhancement of microbial reductive metabolism in post-antibiotic rats. These findings suggested that pectin supplementation accelerated post-antibiotic gut microbiome reconstitution orchestrated with reduced gut redox potential and that the effect of pectin on redox potential was mediated by remodeling of the intestinal microbiota.

Neural control of fluid homeostasis is engaged below 10°C in hibernation.

Thirteen-lined ground squirrels (Ictidomys tridecemlineatus) hibernate for several months each winter without access to water,1 but the mechanisms that maintain fluid homeostasis during hibernation are poorly understood. In torpor, when body temperature (TB) reaches 4°C, squirrels decrease metabolism, slow heart rate, and reduce plasma levels of the antidiuretic hormones arginine vasopressin (AVP) and oxytocin (OXT).1 Squirrels spontaneously undergo interbout arousal (IBA) every 2 weeks, temporarily recovering an active-like metabolism and a TB of 37°C for up to 48 h.1,2 Despite the low levels of AVP and OXT during torpor, profound increases in blood pressure and heart rate during the torpor-IBA transition are not associated with massive fluid loss, suggesting the existence of a mechanism that protects against diuresis at a low TB. Here, we demonstrate that the antidiuretic hormone release pathway is activated by hypothalamic supraoptic nucleus (SON) neurons early in the torpor-arousal transition. SON neuron activity, dense-core vesicle release from the posterior pituitary, and plasma hormone levels all begin to increase before TB reaches 10°C. In vivo fiber photometry of SON neurons from hibernating squirrels, together with RNA sequencing and c-FOS immunohistochemistry, confirms that SON is electrically, transcriptionally, and translationally active to monitor blood osmolality throughout the dynamic torpor-arousal transition. Our work emphasizes the importance of the antidiuretic pathway during the torpor-arousal transition and reveals that the neurophysiological mechanism that coordinates the hormonal response to retain fluid is active at an extremely low TB, which is prohibitive for these processes in non-hibernators.

Electrochemically and Thermally Stable Inorganics-Rich Solid Electrolyte Interphase for Robust Lithium Metal Batteries.

Severe dendrite growth and high-level activity of the lithium metal anode lead to a short life span and poor safety, seriously hindering the practical applications of lithium metal batteries. With a trisalt electrolyte design, an F-/N-containing inorganics-rich solid electrolyte interphase on a lithium anode is constructed, which is electrochemically and thermally stable over long-term cycles and safety abuse conditions. As a result, its Coulombic efficiency can be maintained over 98.98% for 400 cycles. An 85.0% capacity can be retained for coin-type full cells with a 3.14 mAh cm-2 LiNi0.5 Co0.2 Mn0.3 O2 cathode after 200 cycles and 1.0 Ah pouch-type full cells with a 4.0 mAh cm-2 cathode after 72 cycles. During the thermal runaway tests of a cycled 1.0 Ah pouch cell, the onset and triggering temperatures were increased from 70.8 °C and 117.4 °C to 100.6 °C and 153.1 °C, respectively, indicating a greatly enhanced safety performance. This work gives novel insights into electrolyte and interface design, potentially paving the way for high-energy-density, long-life-span, and thermally safe lithium metal batteries.

Lineage divergence at the first TCR-dependent checkpoint: preferential γδ and impaired αß T cell development in nonobese diabetic mice.

The first TCR-dependent checkpoint in the thymus determines αß versus γδ T lineage fate and sets the stage for later T cell differentiation decisions. We had previously shown that early T cells in NOD mice that are unable to rearrange a TCR exhibit a defect in checkpoint enforcement at this stage. To determine if T cell progenitors from wild-type NOD mice also exhibit cell-autonomous defects in development, we investigated their differentiation in the Notch-ligand-presenting OP9-DL1 coculture system, as well as by analysis of T cell development in vivo. Cultured CD4 and CD8 double-negative cells from NOD mice exhibited major defects in the generation of CD4 and CD8 double-positive αß T cells, whereas γδ T cell development from bipotent precursors was enhanced. Limiting dilution and single-cell experiments show that the divergent effects on αß and γδ T cell development did not spring from biased lineage choice but from increased proliferation of γδ T cells and impaired accumulation of αß T lineage double-positive cells. In vivo, NOD early T cell subsets in the thymus also show characteristics indicative of defective ß-selection, and peripheral αß T cells are poorly established in mixed bone marrow chimeras, contrasting with strong γδ T as well as B cell repopulation. Thus, NOD T cell precursors reveal divergent, lineage-specific differentiation abnormalities in vitro and in vivo from the first TCR-dependent developmental choice point, which may have consequences for subsequent lineage decisions and effector functions.

Short-Term Supplementation of Pectin Alters Substrate Dynamics and Modulates Microbial Carbohydrate Metabolism in the Gut of a Pig Model.

The interaction of pectin and gut microbiota plays an important role in maintaining animal and human health, but this interaction is not fully understood. Here, the impact of pectin supplementation on substrate dynamics and gut microbiota (in the terminal ileum and feces) was integrally investigated in a fistula pig model. Our results showed that a pectin-supplemented diet (PEC) decreased the concentrations of starch, cellulose, and butyrate in feces but not in the terminal ileum. Metagenomic sequencing revealed that PEC had a low impact on the ileal microbiota but significantly increased plant polysaccharide-degrading genera (e.g., Bacteroides, Alistipes, and Treponema) in feces. Additionally, CAZyme profiling indicated that PEC reduced GH68 and GH8 for oligosaccharide degradation in the ileal microbiome, while it enriched GH5, GH57, and GH106 for degradation of carbohydrate substrates in feces. Metabolomic analysis confirmed that PEC increased metabolites involved in carbohydrate metabolism including glucuronate and aconitate. Collectively, pectin could promote complex carbohydrate substrate degradation in the hindgut via modulating the gut microbiota.

Supplementation of Schisandrin B in Semen Extender Improves Quality and Oxidation Resistance of Boar Spermatozoa Stored at 4 °C.

During cold storage, boar spermatozoa undergo oxidative stress, which can impair sperm function and fertilizing capacity. The objective of the present study was to assess the effects of Schisandrin B (Sch B) in semen extenders on the quality of boar semen stored at hypothermia. Semen was collected from twelve Duroc boars and diluted in extenders supplemented with different concentrations of Sch B (0 µmol/L, 2.5 µmol/L, 5 µmol/L, 10 µmol/L, 20 µmol/L, and 40 µmol/L). Here, we demonstrated that 10 µmol/L Sch B provided the best effects on motility, plasma membrane integrity, acrosome integrity, sperm normality rate, average movement velocity, wobbility, mitochondrial membrane potential (MMP), and DNA integrity of sperm. The results of Sch B effects on antioxidant factors in boar sperm showed that Sch B significantly elevated the total antioxidant capacity (T-AOC) and markedly decreased the reactive oxygen species (ROS) and malondialdehyde (MDA) content of sperm. The expression of catalase (CAT) and superoxide dismutase (SOD) mRNA was increased, while the expression of glutathione peroxidase (GPx) mRNA demonstrated no change compared to non-treated boar sperm. Compared to the non-treated group, Sch B triggered a decrease in Ca2+/protein kinase A (PKA) and lactic acid content in boar sperm. Similarly, Sch B led to a statistically higher quantitative expression of AWN mRNA and a lower quantitative expression of porcine seminal protein I (PSP-I) and porcine seminal protein II (PSP-II) mRNA. In a further reverse validation test, no significant difference was observed in any of the parameters, including adhesion protein mRNA, calcium content, lactic acid content, PKA, and protein kinase G (PKG) activity after sperm capacitation. In conclusion, the current study indicates the efficient use of Sch B with a 10 µmol/L concentration in the treatment of boar sperm through its anti-apoptosis, antioxidative, and decapacitative mechanisms, suggesting that Sch B is a novel candidate for improving antioxidation and decapacitation factors in sperm in liquid at 4 °C.

Thermoresponsive Electrolytes for Safe Lithium-Metal Batteries.

Exploring advanced strategies in alleviating the thermal runaway of lithium-metal batteries (LMBs) is critically essential. Herein, a novel electrolyte system with thermoresponsive characteristics is designed to largely enhance the thermal safety of 1.0 Ah LMBs. Specifically, vinyl carbonate (VC) with azodiisobutyronitrile is introduced as a thermoresponsive solvent to boost the thermal stability of both the solid electrolyte interphase (SEI) and electrolyte. First, abundant poly(VC) is formed in SEI with thermoresponsive electrolyte, which is more thermally stable against lithium hexafluorophosphate compared to the inorganic components widely acquired in routine electrolyte. This increases the critical temperature for thermal safety (the beginning temperature of obvious self-heating) from 71.5 to 137.4 °C. The remained VC solvents can be polymerized into poly(VC) as the battery temperature abnormally increases. The poly(VC) can not only afford as a barrier to prevent the direct contact between electrodes, but also immobilize the free liquid solvents, thereby reducing the exothermic reactions between electrodes and electrolytes. Consequently, the internal-short-circuit temperature and "ignition point" temperature (the starting temperature of thermal runaway) of LMBs are largely increased from 126.3 and 100.3 °C to 176.5 and 203.6 °C. This work provides novel insights for pursuing thermally stable LMBs with the addition of various thermoresponsive solvents in commercial electrolytes.

Puerol and pueroside derivatives from Pueraria lobata and their anti-inflammatory activity.

Five undescribed puerols and puerosides and three known analogues were obtained from the roots of Pueraria lobata. Their structures were determined by comprehensive analysis of spectroscopic data and chemical methods. Since puerol D and puerol C were racemic compounds, resolved into their enantiomers, and their absolute configurations were determined by experimental and calculated ECD spectra. Six of the isolates were evaluated for their inhibitory activities on NO generation and the expression of inflammatory factors in the LPS-stimulated RAW 264.7 macrophage cells. The results showed that (S)-puerol C, (R)-puerol C, isokuzubutenolide A and kuzubutenolide A significantly decreased the NO production (IC50 values in the range of 16.87-39.95 µM). Meanwhile, (S)-puerol C, isokuzubutenolide A and kuzubutenolide A also reduced the mRNA expression of inflammatory factors (TNF-α, IL-1ß and IL-6).

Fine-scale staging of T cell lineage commitment in adult mouse thymus.

T cell development is marked by the loss of alternative lineage choices accompanying specification and commitment to the T cell lineage. Commitment occurs between the CD4 and CD8 double-negative (DN) 2 and DN3 stages in mouse early T cells. To determine the gene regulatory changes that accompany commitment, we sought to distinguish and characterize the earliest committed wild-type DN adult thymocytes. A transitional cell population, defined by the first downregulation of surface c-Kit expression, was found to have lost the ability to differentiate into dendritic cells and NK cells when cultured without Notch-Delta signals. In the presence of Notch signaling, this subset generates T lineage descendants in an ordered precursor-product relationship between DN2, with the highest levels of surface c-Kit, and c-Kit-low DN3 cells. These earliest committed cells show only a few differences in regulatory gene expression, compared with uncommitted DN2 cells. They have not yet established the full expression of Notch-related and T cell differentiation genes characteristic of DN3 cells before beta selection. Instead, the downregulation of select stem cell and non-T lineage genes appears to be key to the extinction of alternative lineage choices.

To hum or not to hum: Neural transcriptome signature of male courtship vocalization in a teleost fish.

For many animal species, vocal communication is a critical social behavior and often a necessary component of reproductive success. Additionally, vocalizations are often demanding motor acts. Wanting to know whether a specific molecular toolkit might be required for vocalization, we used RNA-sequencing to investigate neural gene expression underlying the performance of an extreme vocal behavior, the courtship hum of the plainfin midshipman fish (Porichthys notatus). Single hums can last up to 2 h and may be repeated throughout an evening of courtship activity. We asked whether vocal behavioral states are associated with specific gene expression signatures in key brain regions that regulate vocalization by comparing transcript expression levels in humming versus non-humming males. We find that the circadian-related genes period3 and Clock are significantly upregulated in the vocal motor nucleus and preoptic area-anterior hypothalamus, respectively, in humming compared with non-humming males, indicating that internal circadian clocks may differ between these divergent behavioral states. In addition, we identify suites of differentially expressed genes related to synaptic transmission, ion channels and transport, neuropeptide and hormone signaling, and metabolism and antioxidant activity that together may support the neural and energetic demands of humming behavior. Comparisons of transcript expression across regions stress regional differences in brain gene expression, while also showing coordinated gene regulation in the vocal motor circuit in preparation for courtship behavior. These results underscore the role of differential gene expression in shifts between behavioral states, in this case neuroendocrine, motor and circadian control of courtship vocalization.

Transwell isolation and difference analysis of capacitated boar sperm proteins based on the iTRAQ technique.

During capacitation, proteins in boar sperm are released to maintain the stability of their own state and membrane structure. No studies have analyzed the differences between retained proteins and released proteins during sperm capacitation. In the present study, a Transwell chamber and polycarbonate membrane were used to separate the proteins of boar sperm and their released proteins. Isotopically labeled relative and absolute quantification (iTRAQ) was used to analyze each compartment protein. A total of 108 differential proteins were identified in the upper and lower chambers of the Transwell, among which 27 were significantly upregulated (p-value≤0.05 and |log2 (fold change)|≥1) and 81 were significantly downregulated (p-value≤0.05 and |log2 (fold change)|≤1). These differential proteins were mainly involved in biological processes (e.g., the regulation of cysteine peptidase activity, transmembrane transportation, ion transportation and ATP synthesis) and major signaling pathways (e.g., glutathione/galactose metabolism, cellular adhesion and PI3K-Akt), and most of them interacted with each other to some extent. In conclusion, retained proteins and released proteins of capacitated sperm were effectively separated using a Transwell chamber, and differential proteins were successfully identified from among the proteins. Bioinformatics analysis suggested that these differential proteins affect sperm capacitation mainly by adjusting sperm energy metabolism, motion characteristics and acrosome membrane status.