How denitrifiers defense ciprofloxacin: Insights from intracellular and extracellular stress response.

Overuse of antibiotics has led to their existence in nitrogen-containing water. The impacts of antibiotics on bio-denitrification and the metabolic response of denitrifiers to antibiotics are unclear. We systematically analyzed the effect of ciprofloxacin (CIP) on bio-denitrification and found that 5 mg/L CIP greatly inhibited denitrification with a model denitrifier (Paracoccus denitrificans). Nitrate reduction decreased by 32.89 % and nitrous oxide emission increased by 75.53 %. The balance analysis of carbon and nitrogen metabolism during denitrification showed that CIP exposure blocked electron transfer and reduced the flow of substrate metabolism used for denitrification. Proteomics results showed that CIP exposure induced denitrifiers to use the pentose phosphate pathway more for substrate metabolism. This caused a substrate preference to generate NADPH to prevent cellular damage rather than NADH for denitrification. Notably, despite denitrifiers having antioxidant defenses, they could not completely prevent oxidative damage caused by CIP exposure. The effect of CIP exposure on denitrifiers after removal of extracellular polymeric substances (EPS) demonstrated that EPS around denitrifiers formed a barrier against CIP. Fluorescence and infrared spectroscopy revealed that the binding effect of proteins in EPS to CIP prevented damage. This study shows that denitrifiers resist antibiotic stress through different intracellular and extracellular defense strategies.

The novel regulator HdrR controls the transcription of the heterodisulfide reductase operon hdrBCA in Methanosarcina barkeri.

Methanogenic archaea play a key role in the global carbon cycle because these microorganisms remineralize organic compounds in various anaerobic environments. The microorganism Methanosarcina barkeri is a metabolically versatile methanogen, which can utilize acetate, methanol, and H2/CO2 to synthesize methane. However, the regulatory mechanisms underlying methanogenesis for different substrates remain unknown. In this study, RNA-seq analysis was used to investigate M. barkeri growth and gene transcription under different substrate regimes. According to the results, M. barkeri showed the best growth under methanol, followed by H2/CO2 and acetate, and these findings corresponded well with the observed variations in genes transcription abundance for different substrates. In addition, we identified a novel regulator, MSBRM_RS03855 (designated as HdrR), which specifically activates the transcription of the heterodisulfide reductase hdrBCA operon in M. barkeri. HdrR was able to bind to the hdrBCA operon promoter to regulate transcription. Furthermore, the structural model analyses revealed a helix-turn-helix domain, which is likely involved in DNA binding. Taken together, HdrR serves as a model to reveal how certain regulatory factors control the expression of key enzymes in the methanogenic pathway.IMPORTANCEThe microorganism Methanosarcina barkeri has a pivotal role in the global carbon cycle and contributes to global temperature homeostasis. The consequences of biological methanogenesis are far-reaching, including impacts on atmospheric methane and CO2 concentrations, agriculture, energy production, waste treatment, and human health. As such, reducing methane emissions is crucial to meeting set climate goals. The methanogenic activity of certain microorganisms can be drastically reduced by inhibiting the transcription of the hdrBCA operon, which encodes heterodisulfide reductases. Here, we provide novel insight into the mechanisms regulating hdrBCA operon transcription in the model methanogen M. barkeri. The results clarified that HdrR serves as a regulator of heterodisulfide reductase hdrBCA operon transcription during methanogenesis, which expands our understanding of the unique regulatory mechanisms that govern methanogenesis. The findings presented in this study can further our understanding of how genetic regulation can effectively reduce the methane emissions caused by methanogens.

Effects of a 10-Week Exercise and Nutritional Intervention with Variable Dietary Carbohydrates and Glycaemic Indices on Substrate Metabolism, Glycogen Storage, and Endurance Performance in Men: A Randomized Controlled Trial.

BACKGROUND: Daily nutrition plays an important role in supporting training adaptions and endurance performance. The objective of this 10-week study was to investigate the consequences of varying carbohydrate consumption and the glycaemic index (GI) together with an endurance training regimen on substrate oxidation, muscle energy storage and endurance performance under free-living conditions. Sixty-five moderately trained healthy men (29 ± 4 years; VO2 peak 55 ± 8 mL min-1 kg-1) were randomized to one of three different nutritional regimes (LOW-GI: 50-60% CHO with ≥ 65% of these CHO with GI < 50 per day, n = 24; HIGH-GI: 50-60% CHO with ≥ 65% CHO with GI > 70 per day, n = 20; LCHF: ≤ 50 g CHO daily, n = 21). Metabolic alterations and performance were assessed at baseline (T0) and after 10 weeks (T10) during a graded exercise treadmill test. Additionally, a 5 km time trial on a 400-m outdoor track was performed and muscle glycogen was measured by magnet resonance spectroscopy. RESULTS: Total fat oxidation expressed as area under the curve (AUC) during the graded exercise test increased in LCHF (1.3 ± 2.4 g min-1 × km h-1, p < 0.001), remained unchanged in LOW-GI (p > 0.05) and decreased in HIGH-GI (- 1.7 ± 1.5 g min-1 × km h-1, p < 0.001). After the intervention, LOW-GI (- 0.4 ± 0.5 mmol L-1 × km h-1, p < 0.001) and LCHF (- 0.8 ± 0.7 mmol L-1 × km h-1, p < 0.001) showed significantly lower AUC of blood lactate concentrations. Peak running speed increased in LOW-GI (T0: 4.3 ± 0.4 vs. T10: 4.5 ± 0.3 m s-1, p < 0.001) and HIGH-GI (T0: 4.4 ± 0.5 vs. T10: 4.6 ± 0.4 m s-1), while no improvement was observed in LCHF. Yet, time trial performance improved significantly in all groups. Muscle glycogen content increased for participants in HIGH-GI (T0: 97.3 ± 18.5 vs. T10: 144.5 ± 39.8 mmol L wet-tissue-1, p = 0.027) and remained unchanged in the LOW-GI and the LCHF group. At the last examination, muscle glycogen concentration was significantly higher in LOW-GI compared to LCHF (p = 0.014). CONCLUSION: Changes in fat oxidation were only present in LCHF, however, lower lactate concentrations in LOW-GI resulted in changes indicating an improved substrate metabolism. Compared to a LCHF diet, changes in peak running speed, and muscle glycogen stores were superior in LOW- and HIGH-GI diets. The low GI diet seems to have an influence on substrate metabolism without compromising performance at higher intensities, suggesting that a high-carbohydrate diet with a low GI is a viable alternative to a LCHF or a high GI diet. TRIAL REGISTRATION: Clinical Trials, NCT05241730. https://clinicaltrials.gov/study/NCT05241730 . Registered 25 January 2021.

[Effects of manipulating lactate dehydrogenase gene on metabolism of HEK-293 and production of human adenovirus].

Reducing lactate accumulation has always been a goal of the mammalian cell biotechnology industry. When animal cells are cultured in vitro, the accumulation of lactate is mainly the combined result of two metabolic pathways. On one hand, glucose generates lactate under the function of lactate dehydrogenase A (LDHA); on the other hand, lactate can be oxidized to pyruvate by LDHB or LDHC and re-enter the TCA cycle. This study comprehensively evaluated the effects of LDH manipulation on the growth, metabolism and human adenovirus (HAdV) production of human embryonic kidney 293 (HEK-293) cells, providing a theoretical basis for engineering the lactate metabolism in mammalian cells. By knocking out ldha gene and overexpression of ldhb and ldhc genes, the metabolic efficiency of HEK-293 cells was effectively improved, and HAdV production was significantly increased. Compared with the control cell, LDH manipulation promoted cell growth, reduced the accumulation of lactate and ammonia, significantly enhanced the efficiency of substrate and energy metabolism of cells, and significantly increased the HAdV production capacity of HEK-293 cells. Among these LDH manipulation measures, ldhc gene overexpression performed the best, with the maximum cell density increased by about 38.7%. The yield of lactate to glucose and ammonia to glutamine decreased by 33.8% and 63.3%, respectively; and HAdV titer increased by at least 16 times. In addition, the ATP production rate, ATP/O2 ratio, ATP/ADP ratio and NADH content of the modified cell lines were increased to varying degrees, and the energy metabolic efficiency was significantly improved.

2'-fucosyllactose alone or combined with resistant starch increases circulating short-chain fatty acids in lean men and men with prediabetes and obesity.

Background: Infusion of short-chain fatty acids (SCFA) to the distal colon beneficially affects human substrate and energy metabolism. Here, we hypothesized that the combination of 2'-fucosyllactose (2'-FL) with resistant starch (RS) increases distal colonic SCFA production and improves metabolic parameters. Methods: In this randomized, crossover study, 10 lean (BMI 20-24.9 kg/m2) and nine men with prediabetes and overweight/obesity (BMI 25-35 kg/m2) were supplemented with either 2'-FL, 2'-FL+RS, or placebo one day before a clinical investigation day (CID). During the CID, blood samples were collected after a overnight fast and after intake of a liquid high-fat mixed meal to determine plasma SCFA (primary outcomes). Secondary outcomes were fasting and postprandial plasma insulin, glucose, free fatty acid (FFA), glucagon-like peptide-1, and peptide YY concentrations. In addition, fecal SCFA and microbiota composition, energy expenditure and substrate oxidation (indirect calorimetry), and breath hydrogen excretion were determined. Results: In lean men, supplementation with 2'-FL increased postprandial plasma acetate (P = 0.017) and fasting H2 excretion (P = 0.041) compared to placebo. Postprandial plasma butyrate concentration increased after 2'-FL and 2'-FL+RS as compared to placebo (P < 0.05) in lean men and men with prediabetes and overweight/obesity. Additionally, 2'-FL+RS decreased fasting and postprandial plasma FFA concentrations compared to placebo (P < 0.05) in lean men. Conclusion: Supplementation of 2'-FL with/without RS the day before investigation increased systemic butyrate concentrations in lean men as well as in men with prediabetes and obesity, while acetate only increased in lean men. The combination of 2'-FL with RS showed a putatively beneficial metabolic effect by lowering plasma FFA in lean men, indicating a phenotype-specific effect. Clinical trial registration: nr. NCT04795804.

Past and present of beta arrestins: A new perspective on insulin secretion and effect.

BACKGROUND: Beta arrestins had been known as intracellular adaptors that uncouple and inactivate the G protein-coupled receptors that they interact with. Their roles as signal initiators for some receptors have recently been recognized. SCOPE OF REVIEW: In this review, we focused on their role in mediating metabolic modulation primarily in relation to insulin signaling. Commenced by the upstream receptor, they seem to act like intracellular hubs that divert the metabolic profile of the cell. The amount of metabolic substrates in circulation and their usage/deposition by tissues are controlled by the contribution of all systems in the organism. This control is enabled by the release of hormones such as insulin, glucagon and glucagon-like peptide-1. Intriguingly, some ligands -either agonists or antagonists-of different classes of receptors have preferential properties mediated by ß arrestins. This is not surprizing considering that substrate supply and usage should parallel physiological function such as hormone release or muscle contraction. MAJOR CONCLUSIONS: Available data indicate that ß arrestins conduct the regulatory role in insulin secretion and action. They may be good candidates to target when the upstream signal demands the function that may compromise the cell. An example is carvedilol that is protective by preventing the stimulatory effects of excessive catecholamines, stimulates mitochondrial function and has preferential clinical outcomes in metabolic disorders.

Longer-Term Effects of the Glycaemic Index on Substrate Metabolism and Performance in Endurance Athletes.

Nutrition has a decisive influence on athletic performance. However, it is not only the nutrient intake during exercise that is important, but the daily diet must also be adapted to the requirements of physical activity in order to optimally promote training adaptations. The goal of prolonged endurance training is to enhance fat oxidation, to maintain aerobic performance at a higher intensity while sparing limited carbohydrate stores. The targeted modification of macronutrient intake is a common method of influencing substrate metabolism, fuel selection, and performance. However, it is not well established whether the glycaemic index of carbohydrates in our daily diet can improve endurance performance by influencing carbohydrate or fat oxidation during training. Therefore, the aim of the following review is to elucidate the possible influence of the glycaemic index on substrate utilization during exercise and to clarify whether the consumption of a long-term high-carbohydrate diet with different glycaemic indices may have an influence on substrate metabolism and endurance performance.

How ß-Cyclodextrin-Functionalized Biochar Enhanced Biodenitrification in Low C/N Conditions via Regulating Substrate Metabolism and Electron Utilization.

Biodenitrification plays a vital role in the remediation of nitrogen-contaminated water. However, influent with a low C/N ratio limits the efficiency of denitrification and causes the accumulation/emission of noxious intermediates. In this study, ß-cyclodextrin-functionalized biochar (BC@ß-CD) was synthesized and applied to promote the denitrification performance of Paracoccus denitrificans when the C/N was only 4, accompanied by increased nitrate reduction efficiency and lower nitrite accumulation and nitrous oxide emission. Transcriptomic and enzymatic activity analyses showed BC@ß-CD enhanced glucose degradation by promoting the activities of glycolysis (EMP), the pentose phosphate pathway (PPP), and the tricarboxylic acid (TCA) cycle. Notably, BC@ß-CD drove a great generation of electron donors by stimulating the TCA cycle, causing a greater supply of substrate metabolism to denitrification. Meanwhile, the promotional effect of BC@ß-CD on oxidative phosphorylation accelerates electron transfer and ATP synthesis. Moreover, the presence of BC@ß-CD increased the intracellular iron level, causing further improved electron utilization in denitrification. BC@ß-CD helped to remove metabolites and induced positive feedback on the metabolism of P. denitrificans. Collectively, these effects elevated the glucose utilization for supporting denitrification from 36.37% to 51.19%. This study reveals the great potential of BC@ß-CD for enhancing denitrification under low C/N conditions and illustrates a potential application approach for ß-CD in wastewater bioremediation.

Extracellular flux analysis in intact cardiac tissue slices-A novel tool to investigate cardiac substrate metabolism in mouse myocardium.

AIM: Cardiac pathologies are accompanied by alterations in substrate metabolism, and extracellular flux analysis is a standard tool to investigate metabolic disturbances, especially in immortalized cell lines. However, preparations of primary cells, such as adult cardiomyocytes require enzymatic dissociation and cultivation affecting metabolism. Therefore, we developed a flux analyzer-based method for the assessment of substrate metabolism in intact vibratome-sliced mouse heart tissue. METHODS: Oxygen consumption rates were determined using a Seahorse XFe24-analyzer and "islet capture plates." We demonstrate that tissue slices are suitable for extracellular flux analysis and metabolize both free fatty acids (FFA) and glucose/glutamine. Functional integrity of tissue slices was proven by optical mapping-based assessment of action potentials. In a proof-of-principle approach, the sensitivity of the method was tested by analyzing substrate metabolism in the remote myocardium after myocardial infarction (I/R). RESULTS: Here, I/R increased uncoupled OCR compared with sham animals indicating a stimulated metabolic capacity. This increase was caused by a higher glucose/glutamine metabolism, whereas FFA oxidation was unchanged. CONCLUSION: In conclusion, we describe a novel method to analyze cardiac substrate metabolism in intact cardiac tissue slices by extracellular flux analysis. The proof-of-principle experiment demonstrated that this approach has a sensitivity allowing the investigation of pathophysiologically relevant disturbances in cardiac substrate metabolism.

Metagenomic analysis of the relationship between the microorganisms and the volatiles' development in the wines during spontaneous fermentation from the eastern foothills of the Ningxia Helan mountains in China.

BACKGROUND: The natural fermentation of multispecies microbial communities is responsible for unique flavors of winery regions of the eastern foothills of the Ningxia Helan Mountains in China. However, the participation of different microorganisms in the metabolic network for the development of important flavor substances is not clearly defined. Microbial population and diversity on different fermentation phases of Ningxia wine were analyzed by metagenomic sequencing approach. RESULTS: Gas chromatography-mass spectrometry and ion chromatography were used to identify flavor components, and 13 esters, 13 alcohols, nine aldehydes and seven ketones were detected in volatile substances with odor activity values > 1, and eight organic acids were detected as important flavor components in young wine. Thus, 52 238 predicted protein-coding genes from 24 genera were identified in the Kyoto Encyclopedia of Genes and Genomes level 2 pathways of global and overview maps, and the genes were primarily involved in amino acid metabolism and carbohydrate metabolism. Major microbial genera (Saccharomyces, Tatumella, Hanseniaspora, Lactobacillus, and Lachancea) were closely related to self-characteristic compound metabolism and further contributed to wine flavor. CONCLUSION: This study clarifies the different metabolic roles of microorganisms in flavor formation during Ningxia wine spontaneous fermentation. Saccharomyces, dominant fungi involved in glycolysis and pyruvate metabolism, produces not only ethanol but also two important precursors, pyruvate and acetyl-CoA, which are necessary for the tricarboxylic acid cycle, fatty acid metabolism, amino acid metabolism, and flavor formation. Lactobacillus and Lachancea, dominant bacteria involved in lactic acid metabolism. Tatumella, dominant bacteria involved in amino acid metabolism, fatty acid metabolism, and acetic acid metabolism to produce esters in the Shizuishan City region samples. These findings provide insights into the use of local functional strains to generate unique flavor formation, as well as improved stability and quality, in wine production. © 2023 Society of Chemical Industry.

Carbohydrate, but not fat, oxidation is reduced during moderate-intensity exercise performed in 33 vs. 18 °C at matched heart rates.

PURPOSE: Exposure to environmental heat stress increases carbohydrate oxidation and extracellular heat shock protein 70 (HSP70) concentrations during endurance exercise at matched absolute, external work rates. However, a reduction in absolute work rate typically occurs when unacclimated endurance athletes train and/or compete in hot environments. We sought to determine the effect of environmental heat stress on carbohydrate oxidation rates and plasma HSP70 expression during exercise at matched heart rates (HR). METHODS: Ten endurance-trained, male cyclists performed two experimental trials in an acute, randomised, counterbalanced cross-over design. Each trial involved a 90-min bout of cycling exercise at 95% of the HR associated with the first ventilatory threshold in either 18 (TEMP) or 33 °C (HEAT), with ~ 60% relative humidity. RESULTS: Mean power output (17 ± 11%, P < 0.001) and whole-body energy expenditure (14 ± 8%, P < 0.001) were significantly lower in HEAT. Whole-body carbohydrate oxidation rates were significantly lower in HEAT (19 ± 11%, P = 0.002), while fat oxidation rates were not different between-trials. The heat stress-induced reduction in carbohydrate oxidation was associated with the observed reduction in power output (r = 0.64, 95% CI, 0.01, 0.91, P = 0.05) and augmented sweat rates (r = 0.85, 95% CI, 0.49, 0.96, P = 0.002). Plasma HSP70 and adrenaline concentrations were not increased with exercise in either environment. CONCLUSION: These data contribute to our understanding of how moderate environmental heat stress is likely to influence substrate oxidation and plasma HSP70 expression in an ecologically-valid model of endurance exercise.

The polyphenol epigallocatechin gallate lowers circulating catecholamine concentrations and alters lipid metabolism during graded exercise in man: a randomized cross-over study.

PURPOSE: Physical exercise is shown to mitigate catecholamine metabolites; however, it is unknown if exercise-induced increases in sympatho-adrenal activity or catecholamine metabolites are influenced by ingestion of specific catechins found within green tea. This study explored the impact of epigallocatechin gallate (EGCG) ingestion on catecholamine metabolism during graded cycle exercise in humans. METHODS: Eight males (22.4 ± 3.3 years, BMI:25.7 ± 2.4 kg.m2) performed a randomised, placebo-controlled, single-blind, cross-over trial after consumption (1450 mg) of either EGCG or placebo (PLAC) and performed graded cycling to volitional exhaustion. Venous bloods were taken at rest, 2 h post-ingestion and after every 3-min stage. Blood variables were analysed for catecholamines, catecholamine metanephrines and metabolic variables at rest, 2 h post-ingestion (POST-ING), peak rate of lipid oxidation (FATpeak), lactate threshold (LT) and peak rate of oxygen consumption (VO2peak). Data were analysed using SPSS (Version 26). RESULTS: Resting catecholamine and metanephrines were similar between trials. Plasma adrenaline (AD) was lower in ECGC treatment group between trials at FATpeak (P < 0.05), LT (P < 0.001) and VO2peak (P < 0.01). Noradrenaline (NA) was lower under EGCG at POST (P < 0.05), FATpeak (P < 0.05), LT (P < 0.01) and VO2peak (P < 0.05) compared to PLAC. Metanephrines, glucose and lactate increased similarly with exercise intensity in both trials. Lipid oxidation rate was 32% lower in EGCG at FATpeak (EGCG 0.33 ± 0.14 vs. PLAC 0.49 ± 0.11 g.min-1, P < 0.05). Cycle time to exhaustion was similar (NS). CONCLUSION: Acute EGCG supplementation reduced circulating catecholamines but not; metanephrine, glucose or lactates, response to graded exercise. Lower circulating catecholamines may explain a lower lipid oxidation rate.

Data on cardiac and vascular functionality in ex vivo and in vivo models following acute administration of trimethylamine N-oxide.

This dataset describes in detail the outcomes of acute trimethylamine N-oxide (TMAO) administration on cardiac, vascular and mitochondrial functionality in ex vivo and in vivo models. The accumulation of TMAO in target tissues was assessed after performing heart perfusion or by incubating aortic tissue in a solution containing TMAO. To evaluate the impact of TMAO on mitochondrial function, the aortic rings and heart homogenates of Wistar rats were incubated in a solution containing [9,10-3H] palmitate (5 µCi/ml) or D-[U-14C] glucose (0.625 µCi/ml) in the presence or absence of TMAO with subsequent measurement of substrate oxidation and uptake. The effects of TMAO on the vascular reactivity of isolated conductance and resistance vessels were tested by measuring their response to acetylcholine and sodium nitroprusside. The impact of elevated TMAO levels on cardiac function and infarct size caused by ischemia-reperfusion injury was evaluated in Langendorff perfused heart model. Normal and forced heart functioning was analyzed by echocardiography in CD-1 mouse acute cardiac stress model induced by isoproterenol (10 µg/mouse) upon single and 7 repeated daily administrations of TMAO (120 mg/kg). The data presented in the manuscript provide valuable information on measurements performed under conditions of acutely elevated TMAO levels in experimental models of cardiac and vascular function and energy metabolism. Furthermore, the data have high reuse potential as they could be applied in the planning of future in vitro, ex vivo, and in vivo studies addressing the molecular mechanisms targeted by elevated levels of TMAO.

The effect of acute manipulation of carbohydrate availability on high intensity running performance, running economy, critical speed, and substrate metabolism in trained Male runners.

Completing selected training sessions with reduced glycogen availability is associated with greater signalling and improved muscle oxidative capacity, although it may impact the overall quality of the session. We examined the effects of low carbohydrate availability on high intensity exercise performance, running economy, critical speed, and substrate metabolism. On two occasions, nine male runners (V̇O2peak 60.3 ± 3.3 mL.kg-1.min-1) completed a glycogen depletion protocol involving 90-min at 75%vV̇O2peak followed by 10 × 1-min at 110% vV̇O2peak. This was followed either by high (HIGH) or low (LOW) carbohydrate intake (>6 g.kg-1.day-1 and <50 g.day-1, respectively) until completion of a performance protocol on day 2 consisting of a series of time-trials (TT) (50m to 3000m) and physiological assessments. There were no differences between LOW and HIGH for any TT distance (mean TT performance times for LOW and HIGH were: 3000m TT 651.7 ± 52.8s and 646.4 ± 52.5s, 1500 m TT 304.0 ± 20.2s and 304.2 ± 22.1s, 400 m TT 67.64 ± 4.2s and 67.3 ± 3.8s, 50 m TT 7.27 ± 0.44s and 7.25 ± 0.45s, respectively, P > 0.05), though some athletes performed better in LOW (n = 5). While fat oxidation in LOW was significantly greater than HIGH (Δ0.32 ± 0.14 g.min-1; P < 0.001 at 14 km.h-1 and Δ0.34 ± 0.12 g.min-1 at 16 km.h-1; P < 0.01), running economy did not differ between trials (P > 0.05). Acute manipulation of carbohydrate availability showed immediate effects on substrate metabolism evidenced by greater fat oxidation without changes in RE. Acute low carbohydrate availability did not affect high intensity running performance across a range of distances.Highlights Acute manipulation of muscle glycogen availability using an exercise and dietary manipulation protocol did not affect subsequent high intensity running performance across a range of running distances.Reduced muscle glycogen resulted in a marked increase in fat oxidation in low glycogen condition but no changes in running economy or critical speed.Individual factors should be considered when prescribing high intensity sessions with restricted carbohydrate availability.

Effects of hypoxic exercise on 24-hour glucose profile and substrate metabolism in overweight and obese men with impaired glucose metabolism.

Hypoxic exercise (HE) may have more pronounced effects on glucose homeostasis than exercise under normoxic conditions (NE), but effects on 24-h glucose profile and substrate utilization remain unclear. We investigated the effects of moderate-intensity HE compared with NE on 24-h glucose profile and substrate metabolism in overweight/obese individuals. Ten overweight/obese men with impaired glucose homeostasis participated in a randomized, single-blind, crossover trial. Participants performed moderate-intensity cycling exercise for 4 consecutive days under mild normobaric hypoxic ([Formula: see text]: 15%) or normoxic ([Formula: see text]: 21%) conditions at similar relative exercise intensity (2 × 30 min/day at 50% of maximal heart rate, with a ∼4-wk washout period. Twenty-four-hour glucose levels and systemic oxygen saturation ([Formula: see text]) were monitored throughout the study. At day 5, plasma metabolites and substrate oxidation were determined during a mixed-meal test under normoxic conditions. [Formula: see text] and absolute workload were lower (both P < 0.001), whereas heart rate was comparable during HE compared with NE. HE did not alter mean 24-h, daytime, and nighttime glucose concentrations, and measures of glycemic variability. However, the HE-induced decrease in [Formula: see text] was positively correlated with HE-induced improvements in mean 24-h (rs = 0.683, P = 0.042) and daytime (rs = 0.783, P = 0.013) glucose concentrations. HE at similar relative exercise intensity reduces [Formula: see text] and has comparable effects on mean 24-h glucose concentration and glycemic variability than NE in overweight/obese men with impaired glucose metabolism. Nevertheless, a more pronounced reduction in [Formula: see text] during HE was associated with lower 24-h glucose concentrations, suggesting that a marked hypoxic stimulus is needed to improve glucose homeostasis.NEW & NOTEWORTHY We demonstrate that hypoxic exercise at similar relative exercise intensity (i.e. lower absolute workload) reduces systemic oxygen saturation ([Formula: see text]) and has comparable effects on mean 24-h glucose concentrations and glycemic variability than normoxic exercise in men with overweight/obesity and impaired glucose metabolism. A more pronounced reduction in [Formula: see text] during hypoxic exercise, however, was associated with lower 24-h and daytime glucose concentrations. Our findings suggest that a marked hypoxic stimulus may improve glucose homeostasis.

Effects of manipulating lactate dehydrogenase gene on metabolism of HEK-293 and production of human adenovirus / 生物工程学报

Reducing lactate accumulation has always been a goal of the mammalian cell biotechnology industry. When animal cells are cultured in vitro, the accumulation of lactate is mainly the combined result of two metabolic pathways. On one hand, glucose generates lactate under the function of lactate dehydrogenase A (LDHA); on the other hand, lactate can be oxidized to pyruvate by LDHB or LDHC and re-enter the TCA cycle. This study comprehensively evaluated the effects of LDH manipulation on the growth, metabolism and human adenovirus (HAdV) production of human embryonic kidney 293 (HEK-293) cells, providing a theoretical basis for engineering the lactate metabolism in mammalian cells. By knocking out ldha gene and overexpression of ldhb and ldhc genes, the metabolic efficiency of HEK-293 cells was effectively improved, and HAdV production was significantly increased. Compared with the control cell, LDH manipulation promoted cell growth, reduced the accumulation of lactate and ammonia, significantly enhanced the efficiency of substrate and energy metabolism of cells, and significantly increased the HAdV production capacity of HEK-293 cells. Among these LDH manipulation measures, ldhc gene overexpression performed the best, with the maximum cell density increased by about 38.7%. The yield of lactate to glucose and ammonia to glutamine decreased by 33.8% and 63.3%, respectively; and HAdV titer increased by at least 16 times. In addition, the ATP production rate, ATP/O2 ratio, ATP/ADP ratio and NADH content of the modified cell lines were increased to varying degrees, and the energy metabolic efficiency was significantly improved.

Effects of Low/Medium-Intensity Exercise on Fat Metabolism after a 6-h Fast.

The effects of fasting and different exercise intensities on lipid metabolism were investigated in 12 male students aged 19.9 ± 1.4 years, with maximal oxygen consumption (VO2max) of 50.33 ± 4.0 mL/kg/min, using a counterbalanced design. Each participant ran on a treadmill at 45% and 65% VO2max continuously for 20 min, followed by running at 85% VO2max for 20 min (or until exhaustion) under a fed or fasted state (6 h). The respiratory exchange ratio (RER), blood glucose (BGLU), blood lactate (BLA), and blood triglyceride (TG) were analyzed during exercise. The results showed that the intensity of exercise did not significantly affect the BGLU and TG in the fed state. The levels of both RER and BLA increased as the intensity of exercise increased from low to high (45, 65, and 85% VO2max), and more energy was converted from fat into glucose at a high intensity of exercise. In the fasted state of 6 h, the BGLU level increased parallel to the intensity of exercise. The RER was close to 1.0 at a high intensity of exercise, indicating that more energy was converted from glycogen. At the intensities of 45 and 65% VO2max, the RER and concentration of TG were both lower in the fasted than in the fed state, showing that a higher percentage of energy comes from fat than in the fed state at 45 and 65% VO2max. When running at 85% VO2max, the BGLU concentration was higher in the fasted than in the fed state, indicating that the liver tissues release more BGLU for energy in the fasted state. Therefore, in the fasted state, running at 45% and 65% of VO2max significantly affects lipid metabolism. On the contrary, the higher RER and BGLU concentrations when running at 85% VO2max revealed no significant difference between the two probes. This study suggests that medium- and low-intensity exercise (45 and 65% VO2max) in the fasted state enhances lipid metabolism.

Energy substrate metabolism and oxidative stress in metabolic cardiomyopathy.

Metabolic cardiomyopathy is an emerging cause of heart failure in patients with obesity, insulin resistance, and diabetes. It is characterized by impaired myocardial metabolic flexibility, intramyocardial triglyceride accumulation, and lipotoxic damage in association with structural and functional alterations of the heart, unrelated to hypertension, coronary artery disease, and other cardiovascular diseases. Oxidative stress plays an important role in the development and progression of metabolic cardiomyopathy. Mitochondria are the most significant sources of reactive oxygen species (ROS) in cardiomyocytes. Disturbances in myocardial substrate metabolism induce mitochondrial adaptation and dysfunction, manifested as a mismatch between mitochondrial fatty acid oxidation and the electron transport chain (ETC) activity, which facilitates ROS production within the ETC components. In addition, non-ETC sources of mitochondrial ROS, such as ß-oxidation of fatty acids, may also produce a considerable quantity of ROS in metabolic cardiomyopathy. Augmented ROS production in cardiomyocytes can induce a variety of effects, including the programming of myocardial energy substrate metabolism, modulation of metabolic inflammation, redox modification of ion channels and transporters, and cardiomyocyte apoptosis, ultimately leading to the structural and functional alterations of the heart. Based on the above mechanistic views, the present review summarizes the current understanding of the mechanisms underlying metabolic cardiomyopathy, focusing on the role of oxidative stress.

Sirtuin 1 aggravates hypertrophic heart failure caused by pressure overload via shifting energy metabolism.

Sirtuin1 (SIRT1) is involved in regulating substrate metabolism in the cardiovascular system. Metabolic homeostasis plays a critical role in hypertrophic heart failure. We hypothesize that cardiac SIRT1 can modulate substrate metabolism during pressure overload-induced heart failure. The inducible cardiomyocyte Sirt1 knockout (icSirt1-/-) and its wild type littermates (Sirt1f/f) C57BL/6J mice were subjected to transverse aortic constriction (TAC) surgery to induce pressure overload. The pressure overload induces upregulation of cardiac SIRT1 in Sirt1f/f but not icSirt1-/- mice. The cardiac contractile dysfunctions caused by TAC-induced pressure overload occurred in Sirt1f/f but not in icSirt1-/- mice. Intriguingly, Sirt1f/f heart showed a drastic reduction in systolic contractility and electric signals during post-TAC surgery, whereas icSirt1-/- heart demonstrated significant resistance to pathological stress by TAC-induced pressure overload as evidenced by no significant changes in systolic contractile functions and electric properties. The targeted proteomics showed that the pressure overload triggered downregulation of the SIRT1-associated IDH2 (isocitrate dehydrogenase 2) that resulted in increased oxidative stress in mitochondria. Moreover, metabolic alterations were observed in Sirt1f/f but not in icSirt1-/- heart in response to TAC-induced pressure overload. Thus, SIRT1 interferes with metabolic homeostasis through mitochondrial IDH2 during pressure overload. Inhibition of SIRT1 activity benefits cardiac functions under pressure overload-related pathological conditions.

Hypermetabolism and Substrate Utilization Rates in Pheochromocytoma and Functional Paraganglioma.

The overproduction of catecholamines in pheochromocytoma/paraganglioma (PPGL) induces a hypermetabolic state. The aim of this study was to evaluate the incidence of a hypermetabolic state and differences in substrate metabolism in consecutive PPGL patients divided by catecholamine phenotype. Resting energy expenditure (REE) and respiratory quotient (RQ) were measured in 108 consecutive PPGL patients and 70 controls by indirect calorimetry. Hypermetabolic state was defined according to the Mifflin St. Jeor Equation as a ratio above 110%. Hypermetabolic state was confirmed in 70% of PPGL patients, regardless of phenotype. Older age, prevalence of diabetes mellitus and arterial hypertension were correlated with hypermetabolic PPGL as compared to normometabolic form. Analysis according to overproduced catecholamine showed differences in VCO2 (p < 0.05) and RQ (p < 0.01) and thus different substate metabolism between phenotypes in hypermetabolic form of PPGL. Lipid utilization was higher in the adrenergic phenotype (p = 0.001) and positively associated with the percentage of REE ratio (R = 0.48, p < 0.001), whereas the noradrenergic phenotype preferentially oxidizes carbohydrates (P = 0.001) and is correlated with the percentage of REE ratio (R = 0.60, p < 0.001). Hypermetabolic state in PPGL is a common finding in both catecholamine phenotypes. Hypermetabolic PPGL patients are older and suffer more from diabetes mellitus and arterial hypertension. Under basal conditions, the noradrenergic type preferentially metabolizes carbohydrates, whereas the adrenergic phenotype preferentially metabolizes lipids.