Subsequent Metabolism Research Articles

Functions of aldolase in lipid synthesis of Schizochytrium sp. by gene disruption to switch carbon metabolism

BackgroundAs a key rate-limiting enzyme in the glycolytic pathway of cells, aldolase affects the distribution of intracellular carbon flux and determines the overall ability of subsequent cell metabolism, which are mainly reported in the medical related researches, but rarely involved microorganisms. In this study, the aldolase gene of Schizochytrium limacinum SR21 (ALDOA) was knocked out to explore the effect of regulating carbon flux on cell growth and lipid synthesis.ResultsThe knockout of ALDOA showed an adverse effect on cell growth and total lipids production, which was decreased by 9.6% and 23.2%, respectively, but helped to improve the synthetic ability of polyunsaturated fatty acids (PUFAs), in which the proportion of docosahexaenoic acid (DHA) increased by 22.9%. Analysis of phospholipomics, real-time quantitative PCR and metabolomics revealed that the knockout of ALDOA weakened the glycolysis pathway and tricarboxylic acid cycle to inhibit cell growth, and lowered the Kennedy pathway to reduce the production of total lipids and the synthesis of phospholipids to affect cell metabolism. Correspondingly, the knockout of ALDOA enhanced the metabolic flux of the pentose phosphate pathway to provide more reducing power for PUFAs accumulation and improved the glycerophosphorylcholine acylation pathway to promote the accumulation of DHA.ConclusionsALDOA knockout redistributes the carbon metabolic flux in cells, by weakening the glycolysis, tricarboxylic acid cycle and glyceride synthesis pathway to inhibit cell growth and total lipid production, and strengthening the pentose phosphate pathway and glycerophosphorylcholine acylation pathway to increase the synthesis of PUFAs and DHA accumulation. This study provides a new idea for identifying the aldolase function in microorganisms and a metabolic strategy to improve DHA accumulation in Schizochytrium.

Activation of Heme Metabolism Promotes Tissue Health After Intraarticular Injury or Surgical Exposure.

Posttraumatic osteoarthritis (PTOA) is a well-recognized public health burden without any disease modifying treatment. This occurs despite noted advances in surgical care in the past 50 years. Mitochondrial oxidative damage pathways initiate PTOA after severe injuries like intraarticular fracture that often require surgery and contribute to PTOA after less severe injuries that may or may not require surgery like meniscal injuries. When considering the mitochondrial and redox environment of the injured joint, we hypothesized that activation of heme metabolism, previously associated with healing in many settings, would cause prototypic mitochondrial reprogramming effects in cartilage ideally suited for use at the time of injury repair. Activation of heme metabolism can be accomplished through the gasotransmitter carbon monoxide (CO), which activates hemeoxygenase-1 (HO1) and subsequent heme metabolism. In this study, we employed unique carbon monoxide (CO)-containing foam (COF) to stimulate heme metabolism and restore chondrocyte oxygen metabolism in vitro and in vivo . Doxycycline-inducible, chondrocyte-specific HO1 overexpressing transgenic mice show similar mitochondrial reprogramming after induction compared to COF. CO is retained at least 24 h after COF injection into stifle joints and induces sustained increases in heme metabolism. Lastly, intraarticular injection of COF causes key redox outcomes without any adverse safety outcomes in rabbit stifle joints ex vivo and in vivo . We propose that activation of heme metabolism is an ideal adjuvant to trauma care that replenishes chondrocyte mitochondrial metabolism and restores redox homeostasis.

Effect of Vitamin D3 on Uterine Morphology and Insulin Signaling in a Polycystic Ovary Syndrome (PCOS) Rat Model

Abstract Polycystic ovary syndrome (PCOS) is a common endocrinopathy in women of reproductive age leading to infertility. Besides reproductive and hormonal disturbances, PCOS is often characterized by vitamin D3 (VD) deficiency. This study aimed to determine the effect of VD on uterine histoarchitecture, the biochemical composition of gland secretions, and the insulin signal transduction pathway using a PCOS rat model. The experiment was conducted on four animal groups (n=8/group): control (C), VD supplemented (VD; 500 IU/day), letrozole-treated (PCOS; 1 mg/kg body weight), and VD-treated PCOS (PCOS+VD) group. Herein, VD supplementation did not improve histomorphometric parameters in the PCOS uterus, whereas clearly influenced sugar composition in uterine gland secretions, restoring their content to that observed in the C group. Furthermore, we found that VD can reduce peripheral and local uterine insulin resistance developed in the PCOS rats via activation of the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway and subsequent glucose metabolism in the uterus. To sum up, the present results suggest a possible beneficial role of VD supplementation for the maintenance of uterus functions in PCOS rats.

Size of lipid emulsion droplets influences metabolism in human CD4+ T cells

ScopeTriglyceride-based lipid emulsions are critical for total parenteral nutrition (TPN), but their long-term use has adverse effects, such as severe liver dysfunction necessitating improved formulations. This study compares the uptake mechanism and intracellular fate of novel glycerol-stabilized nano-sized lipid emulsions with conventional emulsions in CD4+ T cells, focusing on their impact on cellular metabolism. Methods and resultsNanoemulsions were formulated with increased glycerol content. Uptake of emulsions in primary human CD4+ T cells was investigated using different endocytic blockers, then quantified by flow cytometry, and visualized by confocal microscopy. To investigate emulsion intracellular fate, fatty acids in membrane phospholipids were quantified by GC-MS/MS and cellular metabolism was assessed by Seahorse technology. Results show T cells internalize both conventional and nano-sized emulsions using macropinocytosis. Fatty acids from emulsions are stored as neutral lipids in intracellular vesicles and are incorporated into phospholipids of cellular membranes. However, only nanoemulsions additionally use clathrin-mediated endocytosis and deliver fatty acids to mitochondria for increased β-oxidation. ConclusionsSize of lipid emulsion droplets significantly influences their uptake and subsequent metabolism in CD4+ T cells. Our results highlight the potential for improved nutrient utilization with nanoemulsions in TPN formulations possibly leading to less adverse effects.

Insights into the biodegradation process of 2,4,5-trichlorophenoxyacetic acid under anaerobic condition

BackgroundChlorophenoxy compounds represent a group of selective herbicides widely used around the world. Chlorophenoxy herbicides are toxic, chemically stable, and can migrate into groundwater through soil leaching, posing a significant threat to drinking water safety and human health. Chlorophenoxy herbicides in groundwater aquifers are subject to anaerobic processes; however, the pathway and microbiology involved in the attenuation of chlorophenoxy herbicides under anaerobic condition are largely unknown. Here, the anaerobic degradation process of 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), a typical chlorophenoxy herbicide, was investigated.ResultsThe initial 52.5 ± 2.3 μM 2,4,5-T was completely degraded by a sediment-derived microbial consortium, with 3,4-dichlorophenol, 2,5-dichlorophenol, 3-chlorophenol (3-CP) and phenol being identified as the intermediate products. Reductive dechlorination of 3-CP to phenol and the subsequent elimination of phenol were the key transformation steps in the overall degradation process of 2,4,5-T. Amplicon sequencing suggested that Dehalobacter, Sulfuricurvum, Bacteroides, Acetobacterium, and Clostridium sensu stricto 7 might contribute to the transformation of 2,4,5-T to phenol, and Smithella, Syntrophorhabdus, Methanofollis and Methanosaeta likely cooperated to accomplish the complete mineralization of phenol.ConclusionsThis study reported the anaerobic degradation of 2,4,5-T via reductive dechlorination and the subsequent syntrophic metabolization of phenol, an intermediate product transformed from 2,4,5-T. Dehalobacter was identified as the organohalide-respiring population catalyzing the reductive dechlorination reaction. Syntrophorhabdus and methanogenic populations were likely involved in anaerobic phenol oxidation and facilitated the complete mineralization of 2,4,5-T.

Characterization and genome sequence of N,N-dimethylformamide degradation in Paracoccus sulfuroxidans DM175A1-1 isolated from activated sludge of wastewater.

N,N-dimethylformamide (DMF) is an organic solvent with stable chemical properties and high boiling point. Based on its good solubility, DMF is widely used in synthetic textile, leather, electronics, pharmaceutical and pesticide industries. However, the DMF pollutes the environment and does harm to human liver function, kidney function, and nerve function. Herein, an efficient DMF-degrading strain, DM175A1-1, was isolated and identified as Paracoccus sulfuroxidans. This strain can use DMF as the sole source of carbon and nitrogen. Whole-genome sequencing of strain DM175A1-1 revealed that it has a 3.99-Mbp chromosome a 120-kbp plasmid1 and a 40-kbp plasmid2. The chromosome specifically harbors the dmfA1 and dmfA2 essential for the initial steps of DMF degradation. And it also carries the some part of genes facilitating subsequent methylotrophic metabolism and glutathione-dependent pathway. Through further DMF tolerance degradation experiments, DM175A1-1 can tolerate DMF concentrations up to 10,000 mg/L, whereas the majority of Paracoccus strains could only show degradation activity below 1,000 mg/L. And the efficiency of organic nitrogen conversion to NH3-N in DMF can reach 99.0% when the hydraulic retention time (HRT) is controlled at 5 days. Meanwhile, it showed a significant degradation effect at a pharmaceutical enterprise in Zhejiang Province with high concentration of DMF wastewater. This study provides a new strain Paracoccus sulfuroxidans DM175A1-1 which shows a significant influence on DMF degradation, and reveals the characterization on its DMF degradation. It lays the foundation for the application of biological method in the efficient degradation of DMF in industrial wastewater.

Metabolism of baicalin by different microbiota determined by MimiCol.

Substances metabolised by the intestinal microbiota can be used as colon markers and are gaining importance. The flavonoid glycoside baicalin has been described in the literature to be metabolised by the intestinal microbiota. The aim of this work was to investigate how the biotransformation of baicalin to baicalein is related to the intestinal microbiota. For this purpose, stool samples from healthy volunteers with different dietary habits were used. From the pre-cultured stool samples, different standard microbiota were obtained which were used for the subsequent metabolism studies in the in vitro model MimiCol. MimiCol represents the ascending section of the colon, the colon ascendens, in terms of available volume, pH-value, redox potential and bacterial abundance. While during the experiments with added standard microbiota a metabolism of baicalin to baicalein could be detected, this was not the case in a series of experiments without added microbiota. This confirmed the hypothesis that the metabolism of baicalin relies on the bacterial species that are present in the colon. The data collected in the MimiCol therefore support the use of baicalin as a potential marker for the determination of the colon arrival. This can be explained by the fact that baicalin in its native form is poorly absorbed from the gastrointestinal tract. Enzymes of the colonic microbiota, namely β-glucuronidases, hydrolyze baicalin to the aglycone baicalein. The resulting aglycone can be absorbed through the intestinal mucosa and detected in blood plasma. This potentially enables the use of baicalin as a marker to determine the time of arrival in the colon.

What Is the Link between Attention-Deficit/Hyperactivity Disorder (ADHD) and Dyslipidemia in Adults? A German Retrospective Cohort Study.

Background: Alterations in the serum lipid profile have been suspected in many psychiatric disorders, such as schizophrenia and depression. However, studies on lipid status in attention-deficit/hyperactivity disorder (ADHD) are sparse and inconsistent. Methods: Using the nationwide, population-based IQVIA Disease Analyzer database, this retrospective cohort study included 5367 outpatients from general practices in Germany aged ≥18 years with a documented first diagnosis of ADHD between January 2005 and December 2021 and 26,835 propensity score-matched individuals without ADHD. Study outcomes were the first diagnosis of lipid metabolism disorders as a function of ADHD within up to 10 years of the index date. The cumulative 10-year incidence was analyzed using Kaplan-Meier curves and compared using the log-rank test. In addition, univariate Cox regression analyses were performed. Results: In the regression analysis, there was no significant association between ADHD and subsequent lipid metabolism disorders in the total population (HR: 0.94; 95% CI: 0.83-1.08), among women (HR: 1.04; 95% CI: 0.84-1.28), and among men (HR: 0.89; 95% CI: 0.74-1.06). In addition, no significant association was observed in the disease-stratified analyses. Conclusions: The findings of this study indicate that ADHD does not exert an influence on lipid metabolism. However, further investigation is warranted, particularly with respect to pharmacological interventions.

Insight into the fate of tolfenpyrad in tea plant (Camellia sinensis L.) from root uptake

Residual pesticides in agricultural environments, including soil and irrigation water, can be taken up by plants, and thus pose a potential risk to food safety. Although tolfenpyrad has been widely used in tea plantations, limited information is available on its root uptake and fate in tea plants (Camellia sinensis L.). Exploring the mechanisms involved is crucial for understanding the migration and accumulation of tolfenpyrad in tea plants, particularly in the edible parts. In this study, root uptake of tolfenpyrad and its subsequent translocation, distribution, and metabolism in tea seedlings were investigated. The results indicated that the passive transport and apoplastic pathway dominated the root uptake of tolfenpyrad. After uptake, tolfenpyrad distributed predominantly in the cell walls (90.8–92.0 %) of roots, resulting in limited upward translocation in water-soluble fractions through transpirational pull, with translocation factor values far <1 (TFstem/root = 0.115–0.453 and TFleaf/stem = 0.039–0.184). Similar accumulation patterns were observed for the carboxylated metabolite PT-CA as well as hydroxylated metabolite PT-OH. Interestingly, the subcellular distribution of PT-CA in stems was much different from that of the parent tolfenpyrad: PT-CA mainly distributed in the stem cell walls (41.72 %) and cell organelles (56.18 %) at 3 h, then gradually transferred into the cell-soluble fractions (33.07 %) after 120 h. Results from the present study indicated limited upward translocation of tolfenpyrad with its main metabolites to leaves. This finding helps to alleviate concerns about environmental residual tolfenpyrad in tea consumption and provides valuable information for the safety evaluation of tolfenpyrad.

Butyrate metabolism in rumen epithelium affected by host and diet regime through regulating microbiota in a goat model

The rumen is an important organ that enables ruminants to digest nutrients. However, the biological mechanism by which the microbiota and its derived fatty acids regulate rumen development is still unclear. In this study, 18 female Haimen goats were selected and slaughtered at d 30, 60, and 90 of age. Multi-omics analyses (rumen microbial sequencing, host transcriptome sequencing, and rumen epithelial metabolomics) were performed to investigate host–microbe interactions from preweaning to postweaning in a goat model. With increasing age, and after the introduction of solid feed, the increased abundances of Prevotella and Roseburia (P < 0.05) showed positive correlations with volatile fatty acid (VFA) levels and morphological parameters (P < 0.05). Epithelial transcriptomic analysis showed that the expression levels of hub genes, including 3-hydroxy-3-methylglutaryl-CoA synthase isoform 2 (HMGCS2), enoyl-CoA hydratase, short chain 1 (ECHS1), and peroxisome proliferator activated receptor gamma (PPARG), were positively associated with animal phenotype (P < 0.05). These hub genes were mainly correlated to VFA metabolism, oxidative phosphorylation, and the mechanistic target of rapamycin kinase (mTOR) and peroxisome proliferator activated receptor (PPAR) signaling pathways (P < 0.05). Moreover, the primary metabolites in the epithelium changed from glucose preweaning to (R)-3-hydroxybutyric acid (BHBA) and acetoacetic acid (ACAC) postweaning (P < 0.05). Diet and butyrate were the major factors shaping epithelial metabolomics in young ruminants (P < 0.05). Multi-omics analysis showed that the rumen microbiota and VFAs were mainly associated with the epithelial transcriptome, and that alterations in gene expression influenced host metabolism. The “butanoate metabolism” pathway, which transcriptomic and metabolomic analyses identified as being upregulated with age, produces ketones that regulate the “oxidative phosphorylation” pathway, which could provide energy for the development of rumen papillae. Our findings reveal the changes that occur in the rumen microbiota, host transcriptome, and metabolome with age, and validate the role of microbiota-derived VFAs in manipulating host gene expression and subsequent metabolism. This study provides insight into the molecular mechanisms of host–microbe interactions in goats and supplies a theoretical basis and guidance for precise nutritional regulation during the critical time window for rumen development of young ruminants.

Liquid Chromatography-Mass Spectrometry-Based Metabolomics Reveals Dynamic Metabolite Changes during Early Postmortem Aging of Donkey Meat.

Metabolic changes in donkey meat during the early postmortem period have not been previously reported. The LC-MS-based metabolomics technique was conducted to understand the metabolic profiles and identify the key metabolites of donkey meat in the first 48 h postmortem. The pH values showed a decreasing trend followed by an increasing trend. Shear force was the lowest at 4 h and the highest at 24 h (p < 0.05). For the metabolome, some candidate biomarker metabolites were identified, such as adenine, inosine, n-acetylhistidine, citric acid, isocitrate, and malic acid. Predominant metabolic pathways, such as citrate cycle (TCA cycle), alanine, aspartate and glutamate metabolism, and purine metabolism, were affected by aging time. Overabundant n-acetylhistidine was identified in LT, declined at 12 h postmortem aging, and then increased. This may explain the significantly lower pH at 12 h postmortem. Adenine was higher at 4 h postmortem, then declined. Decreased ADP may indicate a fast consumption of ATP and subsequent purine metabolism in donkey meat. The results of this study provided new insights into early postmortem aging of donkey meat quality.

How the elastase-induced rabbit aneurysm heals following flow diverter treatment: a histopathological study.

Fibrin deposition is integral to thrombus formation and wound healing. The role of fibrin deposition and subsequent metabolism following flow diversion for aneurysm treatment remains poorly characterized. This study aimed to evaluate the role of fibrin in early thrombus organization after flow diverter treatment. Thirty-five elastase-induced aneurysms were induced in New Zealand white rabbits and subjected to endoluminal flow diversion treatment. The device-bearing arteries were harvested at 1, 3, and 6 months postimplantation and processed for histopathological examination, including a modified picro-Mallory stain (Carstairs method) to visualize fibrin and platelets, immunohistochemical targeting of smooth muscle actin (SMA), and H&E staining for conventional morphological evaluation. Quantitative analysis of tissue components was carried out using the Orbit Image Analysis software. The samples were also assessed qualitatively to investigate the morphology and location of fibrin and other thrombus components within the intra-aneurysmal thrombi. Statistical analyses were conducted using R software version 4.3.1. Fibrin constituted 27.9% of the thrombus tissue within the aneurysm sac for aneurysms harvested at 1 month, and this rate was significantly lower in the 3-month group (10.2%, p = 0.018). The proportion of blood cells within the sac was also notably higher in the 1-month group compared with other time points. The primary tissue filling the dome at 1 month (14/15, 93%) was an unorganized thrombus primarily composed of fibrin, platelets, and red blood cells. Conversely, aneurysms harvested at 1 month had the lowest collagen level (25.6%). However, collagen became the dominant tissue component within the aneurysm sac, accounting for 71.8% of tissue in the 3-month group (p = 0.007). There were no differences observed among the examined components between the 3-month and 6-month groups. On qualitative analysis, collagen-producing SMA-positive myofibroblasts were located near or in between fibrin molecules. Healed aneurysms exhibited myofibroblasts, collagen, and a well-organized fibrin network on the aneurysm neck. In contrast, unhealed aneurysms displayed a poorly organized fibrin network with scattered myofibroblasts at the neck area. These findings indicate that fibrin plays a foundational role in the gradual occlusion of aneurysms after flow diverter treatment. Endovascular approaches that enhance fibrin accumulation could potentially improve aneurysm occlusion rates. Further research is needed to establish the precise role of fibrin in aneurysm occlusion.

CX-4945 (Silmitasertib) Induces Cell Death by Impairing Lysosomal Utilization in KRAS Mutant Cholangiocarcinoma Cell Lines.

Macropinocytosis is a non-selective form of endocytosis that facilitates the uptake of extracellular substances, such as nutrients and macromolecules, into the cells. In KRAS-driven cancers, including pancreatic ductal adenocarcinoma, macropinocytosis and subsequent lysosomal utilization are known to be enhanced to overcome metabolic stress. In this study, we investigated the role of Casein Kinase 2 (CK2) inhibition in macropinocytosis and subsequent metabolic processes in KRAS mutant cholangiocarcinoma (CCA) cell lines. The bovine serum albumin (BSA) uptake indicating macropinocytosis was performed by flow cytometry using the HuCCT1 KRAS mutant CCA cell line. To validate macropinosome, the Rab7 and LAMP2 were labeled and analyzed via immunocytochemistry and western blot. The CX-4945 (Silmitasertib), CK2 inhibitor, was used to investigate the role of CK2 in macropinocytosis and subsequent lysosomal metabolism. The TFK-1, a KRAS wild-type CCA cell line, showed only apoptotic morphological changes. However, the HuCCT1 cell line showed macropinocytosis. Although CX-4945 induced morphological changes accompanied by the accumulation of intracellular vacuoles and cell death, the level of macropinocytosis did not change. These intracellular vacuoles were identified as late macropinosomes, representing Rab7+ vesicles before fusion with lysosomes. In addition, CX-4945 suppressed LAMP2 expression following the inhibition of the Akt-mTOR signaling pathway, which interrupts mature macropinosome and lysosomal metabolic utilization. Macropinocytosis is used as an energy source in the KRAS mutant CCA cell line HuCCT1. The inhibition of CK2 by CX-4945 leads to cell death in HuCCT1 cells through alteration of the lysosome-dependent metabolism.

Missing information on the link between cell death mechanisms and good eating quality in cultured myofoods

BackgroundCommercial interest in the production of cultured myoproducts (cultured “meat”) has grown in a broad range of companies. As has been shown with plant-based meat alternatives, consumer acceptance of such products will depend on their eating quality, as well as cost, ethical and sustainability issues. Scope and approachPrincipal factors in the sensory quality of conventionally-produced meat (texture, colour, flavour, juiciness) are very much affected by post-mortem metabolism in muscle tissue. However, there is a lack of information on how cell death and subsequent metabolism may affect the eating quality of cultured myoproducts. Key finding/hypothesisThe main mechanisms of cell death in cell culture are apoptosis, necrosis and autophagy. Knowledge of the conditions and pathways inducing these and other mechanisms of cell death provides a conceptual framework of how cell harvesting conditions could be used to manipulate post-mortem metabolism in cultured myoproducts. Because the structure, composition and protein isoforms present will differ in conventional meat versus cultured products, a key question will be whether the well-understood post-mortem metabolic processes in the conversion of muscle to meat are desirable, or should be altered, to achieve acceptable sensory qualities of cultured myoproducts. ConclusionPublished information on post-mortem metabolism in cultured muscle cells is entirely lacking. Although post-mortem metabolism will differ with specific cell line and bioreactor conditions, general studies in the area should provide important tools in manipulating the end-product quality of cultured myofoods.

Bile Salt Hydrolase Activity-Based Probes for Monitoring Gut Microbial Bile Acid Metabolism.

Bile acids are bioactive metabolites that are biotransformed into secondary bile acids by the gut microbiota, a vast consortium of microbes that inhabit the intestines. The first step in intestinal secondary bile acid metabolism is carried out by a critical enzyme, bile salt hydrolase (BSH), that catalyzes the gateway reaction that precedes all subsequent microbial metabolism of these important metabolites. As gut microbial metabolic activity is difficult to probe due to the complex nature of the gut microbiome, approaches are needed to profile gut microbiota-associated enzymes such as BSH. Here, we develop a panel of BSH activity-based probes (ABPs) to determine how changes in diurnal rhythmicity of gut microbiota-associated metabolism affects BSH activity and substrate preference. This panel of covalent probes enables determination of BSH activity and substrate specificity from multiple gut anerobic bacteria derived from the human and mouse gut microbiome. We found that both gut microbiota-associated BSH activity and substrate preference is rhythmic, likely due to feeding patterns of the mice. These results indicate that this ABP-based approach can be used to profile changes in BSH activity in physiological and disease states that are regulated by circadian rhythms.

Characteristic substance analysis and rapid detection of bacteria spores in cooked meat products by surface enhanced Raman scattering based on Ag@AuNP array substrate

BackgroundBacterial spores are the main potential hazard in medium- and high-temperature sterilized meat products, and their germination and subsequent reproduction and metabolism can lead to food spoilage. Moreover, the spores of some species pose a health and safety threat to consumers. The rapid detection, prevention, and control of bacterial spores has always been a scientific problem and a major challenge for the medium and high-temperature meat industry. Early and sensitive identification of spores in meat products is a decisive factor in contributing to consumer health and safety. ResultsIn this study, we developed a novel and stable Ag@AuNP array substrate by using a two-step synthesis approach and a liquid-interface self-assembly method that can directly detect bacterial spores in actual meat product samples without the need for additional in vitro bacterial culture. The results indicate that the Ag@AuNP array substrate exhibits high reproducibility and Raman enhancement effects (1.35 × 105). The differentiation in the Surface enhanced Raman scattering (SERS) spectra of five bacterial spores primarily arises from proteins in the spore coat and inner membrane, peptidoglycan of cortex, and Ca2⁺-DPA within the spore core. The correct recognition rate of linear discriminant analysis for spores in the meat product matrix can reach 100 %. The average recovery accuracy of the SERS quantitative model was at around 101.77 %, and the limit of detection can reach below 10 CFU/mL. SignificanceIt provides a promising technological strategy for the characteristic substance analysis and timely monitoring of spores in meat products.

Substituting carbohydrate at lunch for added protein increases fat oxidation during subsequent exercise in healthy males.

How pre-exercise meal composition influences metabolic and health responses to exercise later in the day is currently unclear. Examine the effects of substituting carbohydrate for protein at lunch on subsequent exercise metabolism, appetite, and energy intake. Twelve healthy males completed three trials in randomized, counterbalanced order. Following a standardized breakfast (779 ± 66 kcal; ∼08:15), participants consumed a lunch (1186 ± 140 kcal; ∼13:15) containing either 0.2 g·kg-1 carbohydrate and ∼2 g·kg-1 protein (LO-CARB), 2 g·kg-1 carbohydrate and ∼0.4 g·kg-1 protein (HI-CARB), or fasted (FAST). Participants later cycled at ∼60% V̇O2peak for 1 h (∼16:15) and post-exercise ad-libitum energy intake was measured (∼18:30). Substrate oxidation, subjective appetite, and plasma concentrations of glucose, insulin, non-esterified fatty acids (NEFA), peptide YY (PYY), glucagon-like peptide-1 (GLP-1), and acylated ghrelin (AG) were measured for 5 h post-lunch. Fat oxidation was greater during FAST (+11.66 ± 6.63 g) and LO-CARB (+8.00 ± 3.83 g) than HI-CARB (p < 0.001), with FAST greater than LO-CARB (+3.67 ± 5.07 g; p < 0.05). NEFA were lowest in HI-CARB and highest in FAST, with insulin demonstrating the inverse response (all p < 0.01). PYY and GLP-1 demonstrated a stepwise pattern, with LO-CARB greatest and FAST lowest (all p < 0.01). AG was lower during HI-CARB and LO-CARB versus FAST (p < 0.01). Energy intake in LO-CARB was lower than FAST (-383 ± 233 kcal; p < 0.001) and HI-CARB (-313 ± 284 kcal; p < 0.001). Substituting carbohydrate for protein in a pre-exercise lunch increased fat oxidation, suppressed subjective and hormonal appetite, and reduced post-exercise energy intake.

Metabolite characterisation of the peptide–drug conjugate LN005 in liver S9s by UHPLC-Orbitrap-HRMS

LN005 is a peptide–drug conjugate (PDC) targeting glucose-regulated protein 78 (GRP78) to treat several types of cancer, such as breast, colon, and prostate cancer. As a new drug modality, understanding its metabolism and elimination pathways will help us to have a whole picture of it. Currently, there are no metabolic studies on LN005; therefore, this study aimed to investigate the metabolism of LN005, clarify its metabolic profile in the liver S9s of different species, and identify the major metabolic pathways and differences between species. The incubation samples were measured by ultra-high performance liquid chromatography combined with orbitrap tandem mass spectrometry (UHPLC-Orbitrap-HRMS). The results showed that LN005 was metabolised by liver S9s, and four metabolites were identified. The main metabolic pathway of LN005 in liver S9s was oxidative deamination to ketone or hydrolysis. Similar metabolic profiles were observed in mouse, rat, dog, monkey, and human liver S9s, indicating no differences between these four animal species and humans. This study provides information for the structural modification and optimisation of LN005 and affords a reference for subsequent animal experiments and human metabolism of other PDCs.

240 Prolonged Maternofetal Inflammation in the Pregnant Ewe at Mid-Gestation Increases Subsequent Fetal Inflammatory Tone and Disrupts Skeletal Muscle Growth and Glucose Metabolism Near Term

Abstract Chronic maternofetal inflammation in late gestation (i.e., peak fetal growth) causes fetal intrauterine growth restriction (IUGR) in sheep, which was associated with deficits in growth, body composition, and metabolic function before and after birth. However, the impact of sustained maternofetal inflammation during mid-gestation (i.e., peak placental growth and development) has not been explored. Therefore, our objective was to determine the effects of sustained maternofetal inflammation during this critical window on subsequent fetal inflammatory tone, metabolism, and myoblast (i.e., muscle stem cell) function. Pregnant ewes were injected every 3rd day from the 50th to 65th d of gestational age (dGA; term = 150 dGA) with saline (control; n = 12) or bacterial lipopolysaccharide endotoxin to induce maternofetal inflammation and IUGR (MI-IUGR; n = 12). Fetuses were surgically catheterized on dGA 118 to collect blood samples, and a hyperinsulinemic-euglycemic clamp (HEC) was performed on dGA 123. Fetuses were necropsied on dGA 125 to collect semitendinosus muscle samples. Fetal plasma TNFα concentrations tended to be greater (P = 0.07) for MI-IUGR fetuses (440 ± 45 pg/m l) than for controls (348 ± 24 pg/mL). Fetal blood O2 partial pressures were 12% less (P &amp;lt; 0.05) and maternofetal O2 gradients tended to be 11% greater (P = 0.08) for MI-IUGR fetuses than for controls. Hindlimb-specific glucose uptake rates did not differ between fetal groups, but hindlimb-specific glucose oxidation rates were reduced (P &amp;lt; 0.05) by 15% under basal conditions and by 27% under hyperinsulinemic conditions for MI-IUGR fetuses compared with controls. Semitendinosus glycogen content and β2 adrenoceptor content did not differ between fetal groups, but semitendinosus content of the TNFα receptor, TNFR1, was 41% greater (P &amp;lt; 0.05) for MI-IUGR fetuses than for controls. Immunohistochemistry revealed 18% fewer (P &amp;lt; 0.05) proliferating myoblasts (pax7+/Ki67+) and 38% fewer (P &amp;lt; 0.05) differentiated myoblasts (myogenin+) in semitendinosus from MI-IUGR fetuses compared with controls, which coincided with 13% smaller (P &amp;lt; 0.05) muscle fibers. From these findings, we conclude that sustained maternofetal inflammation during peak placental development at mid-gestation resulted in enhanced fetal inflammatory tone near term, which in turn impaired myoblast function, muscle fiber hypertrophy, and muscle glucose metabolism.

Metabolic reprograming and increased inflammation by cadmium exposure following early-life respiratory syncytial virus infection-the involvement of protein S-palmitoylation.

Early-life respiratory syncytial virus (RSV) infection (eRSV) is one of the leading causes of serious pulmonary disease in children. eRSV is associated with higher risk of developing asthma and compromised lung function later in life. Cadmium (Cd) is a toxic metal, widely present in the environment and in food. We recently showed that eRSV re-programs metabolism and potentiates Cd toxicity in the lung, and our transcriptome-metabolome-wide study showed strong associations between S-palmitoyl transferase expression and Cd-stimulated lung inflammation and fibrosis signaling. Limited information is available on the mechanism by which eRSV re-programs metabolism and potentiates Cd toxicity in the lung. In the current study, we used a mouse model to examine the role of protein S-palmitoylation (Pr-S-Pal) in low dose Cd-elevated lung metabolic disruption and inflammation following eRSV. Mice exposed to eRSV were later treated with Cd (3.3 mg CdCl2/L) in drinking water for 6 weeks (RSV+Cd). The role of Pr-S-Pal was studied using a palmitoyl transferase inhibitor, 2-bromopalmitate (BP, 10 µM). Inflammatory marker analysis showed that cytokines, chemokines and inflammatory cells were highest in the RSV+Cd group, and BP decreased inflammatory markers. Lung metabolomics analysis showed that pathways including phenylalanine, tyrosine and tryptophan, phosphatidylinositol and sphingolipid were altered across treatments. BP antagonized metabolic disruption of sphingolipid and glycosaminoglycan metabolism by RSV+Cd, consistent with BP effect on inflammatory markers. This study shows that Cd exposure following eRSV has a significant impact on subsequent inflammatory response and lung metabolism, which is mediated by Pr-S-Pal, and warrants future research for a therapeutic target.