Phosphofructokinase Activity Research Articles

Type 2 polysaccharide storage myopathy in Quarter Horses is a novel glycogen storage disease causing exertional rhabdomyolysis.

Both type 1 (PSSM1) and type 2 polysaccharide storage myopathy (PSSM2) are characterised by aggregates of abnormal polysaccharide in skeletal muscle. Whereas the genetic basis for PSSM1 is known (R309H GYS1), the cause of PSSM2 in Quarter Horses (PSSM2-QH) is unknown and glycogen concentrations not defined. To characterise the histopathological and biochemical features of PSSM2-QH and determine if an associated monogenic variant exists in genes known to cause glycogenosis. Retrospective case control. Sixty-four PSSM2-QH, 30 PSSM1-QH and 185 control-QH were identified from a biopsy repository and clinical data, histopathology scores (0-3), glycogen concentrations and selected glycolytic enzyme activities compared. Coding sequences of 12 genes associated with muscle glycogenoses were identified from whole genome sequences and compared between seven PSSM2-QH and five control-QH. Exertional rhabdomyolysis in PSSM2-QH occurred predominantly in barrel racing and working cow/roping performance types and improved with regular exercise and a low starch/fat-supplemented diet. Histopathological scores, including the amount of amylase-resistant polysaccharide (PSSM2-QH 1.4 ± 0.6, PSSM1-QH 2.1 ± 0.3, control-QH 0 ± 0, p < 0.001), and glycogen concentrations (PSSM2-QH 129 ± 62, PSSM1-QH 175 ± 9, control-QH 80 ± 27 mmol/kg, p < 0.0001) were intermediate in PSSM2-QH with significant differences among groups. In PSSM2-QH, abnormal polysaccharide had a less filamentous ultrastructure than PSSM1-QH and phosphorylase and phosphofructokinase activities were normal. Seventeen of 30 PSSM2-QH with available pedigrees descended from one of three stallions within four generations. Of the 29 predicted high or moderate impact genetic variants identified in candidate genes, none were present in only PSSM2-QH and absent in control-QH. Analyses of PSSM2-QH and PSSM1-QH were performed on shipped samples, controls on frozen samples. PSSM2-QH is a novel glycogen storage disorder that is not the result of a mutation in genes currently known to cause muscle glycogenoses in other species.

TGF-β3 induces lactate production in Sertoli cell through inhibiting the Notch pathway.

In the testis, spermatocytes and spermatids rely on lactate produced by Sertoli cells (SCs) as energy source. Transforming growth factor-beta 3 (TGF-β3) is one of the generally accepted paracrine regulatory factors of SC-created blood-testis barrier (BTB), yet its role in SC glycolysis and lactate production still remains unclear. To investigate the effect of TGF-β3 on glycolysis and lactate production in SCs and determine the role of lethal giant larvae 2 (Lgl2) and Notch signaling activity during this process. Primary cultured rat SCs and TM4 cells were treated with different concentrations of TGF-β3. In some experiments, cells were transfected with siRNA specifically targeting Lgl2 and then treated with TGF-β3 or N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester. Lactate concentration, glucose and glutamine (Gln) consumption in the culture medium, activity of phosphofructokinase (PFK), lactate dehydrogenase (LDH), and glutaminase (Gls), ATP level, oxygen consumption, extracellular acidification, and mitochondrial respiration complex activity were detected using commercial kits. The protein level of Lgl2, LDH, monocarboxylate transporter 4 (MCT4), and activity of Akt, ERK, p38 MAPK, and Notch pathway were detected by Western blot. The stage-specific expression of Jagged1 was examined by immunohistochemistry (IHC) and qPCR after laser capture microdissection. Spermatogenesis in rat testis injected with recombinant Jagged1 (re-Jagged1) was observed by HE staining, and lactate concentration in testis lysate was measured at a different day point after re-Jagged1 treatment. Significant enhancement of lactate concentration was detected in a culture medium of both primary SCs and TM4 cells treated with TGF-β3 at 3 or 5ng/ml. Besides, other parameters of glycolysis, that is, glucose and Gln consumption, enzyme activity of PFK, LDH, and Gls displayed different levels of increment in primary SCs and TM4 cells after TGF-β3 treatment. Mitochondria respiration of SCs was shown to decrease in response to TGF-β3. Lgl2, MCT4, activity of ERK, and p38 MAPK were up-regulated, whereas Akt and Notch pathway activity were inhibited by TGF-β3. Silencing of Lgl2 in SCs affected lactate production and attenuated the previous effects of TGF-β3 on SC glycolysis except for Gln consumption, Gls activity, and activity of Akt, ERK, and p38. N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT) treatment in SCs antagonized glycolysis suppression caused by Lgl2-silencing. In vivo analysis revealed a stage-specific expression of Jagged1 in contrary with TGF-β3. Activating Notch signaling by re-Jagged1 resulted in restorable hypospermatogenesis and lowered lactate level in rat testis. TGF-β3 induces lactate production in SC through up-regulating Lgl2, which weakened the Notch signaling activity and intensified glycolysis in SCs. Thus, besides the known function of TGF-β3 as the BTB regulator, TGF-β3-Lgl2-Notch may be considered an important pathway controlling SC glycolysis and spermatogenesis.

Hypoxia alters glucose and lipid metabolisms in golden pompano (Trachinotus blochii)

Sufficient dissolved oxygen (DO) is essential for most teleosts. However, high-density aquaculture and fluctuating environmental factors can result in hypoxic stress in golden pompano (Trachinotus blochii). In this study, physiological, biochemical and transcript indicators of metabolism and apoptosis were investigated in T. blochii under hypoxic conditions (1.9 ± 0.2 mg/L) for 12 h. Increases in the content of lactic acid (LD) and the activity of lactate dehydrogenase (LDH), pyruvic acid (PA), phosphofructokinase (PFKA), and hexokinase (HK) suggested the enhancement of anaerobic glycolysis under hypoxic stress. Furthermore, significant increases in triglycerides (TGs) and lipoprotein lipase (LPL) in the liver under hypoxic stress for 6 h and a high level of carnitine palmitoyltransferase-1 (CPT-1) under hypoxic stress for 12 h indicated activation of lipid utilization. Additionally, glycolysis/gluconeogenesis, sphingolipid metabolism, pentose phosphate, HIF-1, and p53 signaling pathways were activated in response to hypoxia. Eleven DEGs involved in these pathways were identified. The relative expressions of HIF-1α (Hypoxia Inducible Factor 1 Subunit Alpha), PGM (phosphoglucomutase), PFKA (6-phosphofructokinase 1), HK (hexokinase), SMPD3 (sphingomyelin phosphodiesterase 3), SPT (serine palmitoyltransferase), and EPO (erythropoietin) were up-regulated under hypoxic conditions. Additionally, the relative expressions of RGN (gluconolactonase), CCNG2 (cyclin G2), CERS1 (sphingoid base N-stearoyltransferase 1), and IGFBP3 (insulin-like growth factor-binding protein 3) were down-regulated. This research reveals metabolic changes and cell apoptosis activation in T. blochii under hypoxic conditions, providing new insight into metabolic responses to hypoxia in teleosts.

Reactive oxygen species-mediated oxidative stress accelerates glycolysis via activation of the CaMKKβ/AMPK pathway in the yak longissimus dorsi postmortem.

Adenosine monophosphate-activated protein kinase (AMPK) is instrumental in the initiation of early postmortem glycolysis and the advent of pale, soft, and exudative (PSE) meat when cellular energy is altered. However, conflicting studies show that AMPK activation without corresponding energy level changes in PSE meat challenges this long-held notion. Here, we examined the effects of reactive oxygen species (ROS)-mediated oxidative stress on AMPK activation in the context of glycolysis, protein solubility, and water-holding capacity (WHC) in the postmortem yak longissimus dorsi (LD) muscle. Further, we explored the mechanisms underlying these effects. Hydrogen peroxide (H2 O2 ) significantly augmented the degree of oxidative stress, increasing the production of ROS and malondialdehyde excessive production and reducing the activity of the anti-oxidants superoxide dismutase and glutathione peroxidase. In turn, oxidative stress dramatically promoted AMPK activation and glycolysis by increasing glycogen depletion and promoting hexokinase and phosphofructokinase activity. Subsequently, lactic acid accumulation increased, leading to a rapid decline in pH, which aggravated protein solubility degree and centrifugal loss in the early postmortem yak LD muscle. Importantly, these changes caused by oxidative stress were eliminated by the AMPK inhibitor. Mechanistically, oxidative stress elevated calcium ion (Ca2+ ) levels, which mobilized calcium/calmodulin-dependent protein kinase β (CaMKKβ) and AMPK. Rescue experiments confirmed that the increases were attenuated using Ca2+ and CaMKKβ chelators, respectively. These results indicated that oxidative stress caused by ROS hastened early-stage postmortem glycolysis and reduced the WHC of yak meat. These effects were likely mediated by the alternative and energy-independent CaMKKβ/AMPK signaling pathway. © 2022 Society of Chemical Industry.

High fat high fructose diet induces mild oxidative stress and reorganizes intermediary metabolism in male mouse liver: Alpha-ketoglutarate effects

BackgroundDiets rich in fats and/or carbohydrates are used to study obesity and related metabolic complications. We studied the effects of a high fat high fructose diet (HFFD) on intermediary metabolism and the development of oxidative stress in mouse liver and tested the ability of alpha-ketoglutarate to prevent HFFD-induced changes. MethodsMale mice were fed a standard diet (10% kcal fat) or HFFD (45% kcal fat, 15% kcal fructose) with or without addition of 1% alpha-ketoglutarate (AKG) in drinking water for 8 weeks. ResultsThe HFFD had no effect on body mass but activated fructolysis and glycolysis and induced inflammation and oxidative stress with a concomitant increase in activity of antioxidant enzymes in the mouse liver. HFFD-fed mice also showed lower mRNA levels of pyruvate dehydrogenase kinase 4 (PDK4) and slightly increased intensity of mitochondrial respiration in liver compared to mice on the standard diet. No significant effects of HFFD on transcription of PDK2 and PGC1α, a peroxisome proliferator-activated receptor co-activator-1α, or protein levels of p-AMPK, an active form of AMP-activated protein kinase, were found. The addition of AKG to HFFD decreased oxidized glutathione levels, did not affect levels of lipid peroxides and PDK4 transcripts but increased activities of hexokinase and phosphofructokinase in mouse liver. ConclusionsSupplementation with AKG had weak modulating effects on HFFD-induced oxidative stress and changes in energetics in mouse liver. General significanceOur research expands the understanding of diet-induced metabolic switching and elucidates further roles of alpha-ketoglutarate as a metabolic regulator.

Ultrasonic treatment decreases Lyophyllum decastes fruiting body browning and affects energy metabolism

Lyophyllum decastes is a common mushroom that is prone to browning during prolonged storage. In this study, the effects of ultrasonic treatment on metabolic gene expression, enzyme activity, and metabolic compounds related to L. decastes browning were investigated. Treatment of the fruiting body at 35 kHz and 300 W for 10 min reduced the browning index of L. decastes by 21.0 % and increased the L* value by 11.1 %. Ultrasonic treatment of the fruiting body resulted in higher levels of total phenols, flavonoids, and 9 kinds of amino acid with catalase (CAT) and peroxidase (POD) activities maintained at high levels. Higher cytochrome c oxidase (CCO), succinate dehydrogenase (SDH), phosphofructokinase (PFK), and pyruvate kinase (PK) activities may be ascribed to increased antioxidant capacity. Moreover, ultrasonication retained higher adenosine triphosphate (ATP) concentrations with an increased energy charge, while there were lower levels of adenosine diphosphate (ADP) and reduced and oxidized nicotinamide adenine dinucleotide (NADH and NAD+), respectively. Meanwhile, lower lignin contents were observed, along with retarded polyphenol oxidase (PPO) and lipoxygenase (LOX) activities. Lower PPO activity reduced the fruiting body enzymatic browning rate through decreased expression of LdPpo1, LdPpo2, and LdPpo3 during storage at 4 °C for 16 days. This activity may be used to determine the effectiveness of ultrasonication.

Inhibition by Ascorbate among Phosphofructokinase-1, Aldolase, Enolase, and Lactate Dehydrogenase in Rabbit Muscle

Background: These studies examined mutual protective relationships among rabbit muscle aldolase, enolase, phosphofructokinase-1 (PFK-1) and LDH from inhibitions by ascorbate (AA). It was proposed earlier that specific inhibitions of PFK-1 and LDH by AA faciltated glycogen storage in resting muscle by inhibiting glycolysis. Materials and Methods: The L-ascorbate (AA), L-ascorbyl dibutyrate (AADB), L-ascorbyl dipalmitate (AADP), L-ascorbyl palmitate (AAP), and L- ascorbyl stearate (AS) are shown in Figure 1 and were obtained from TCI and Alfa Aesar. Unless otherwise stated, all enzymes come from rabbit and all experimental temperatures were 25°C, pH 8.0. Results: Rabbit muscle enolase was examined for its protective effect on other rabbit muscle glycolytic enzymes against inhibitions by ascorbate (AA) and some AA-faty acid derivatives. The IC50 values of enolase by ascorbate (AA) and IC50 values of AA-fatty acid derivatives were compared to estimate inhibition potency. For example, ascorbyl dipalmitate (AADP) was 156 times more inhibitory to enolase than AA. It was previously shown that rabbit muscle aldolase prevented LDH activity loses due to AA inhibition and prevented PFK-1 activity losses due both to dilution and AA inhibition; enolase was found to have the same effects as aldolase. Additionally, PFK-1 prevented enolase and LDH inhibitions by AA. LDH did not prevent enolase or PFK-1 from inhibition by AA. LDH did stimulate enolase activity but not PFK-1 activity. Conclusion: The results suggest that interactions among glycolytic enzyme serve to mutually protect one another from activity losses. The inhibition properties of the AA-fatty acid derivatives are discussed in relation to their possible roles in cancer and diabetes.

Phosphorylation and acetylation of glycolytic enzymes cooperatively regulate their activity and lamb meat quality

This study examined cooperative regulation of phosphorylation and acetylation of glycolytic enzymes on their activity and lamb meat quality. Muscle samples were divided into two groups (fast and slow) according to their glycolysis rate as defined by pH decline rate from 1 h to 1 d postmortem. In slow glycolysis rate group, the activity of hexokinase (HK), phosphofructokinase (PFK) and pyruvate kinase (PK) was lower and meat sample showed lower a*, higher shear force and cooking loss. The acetylation and phosphorylation of HK were positively correlated with HK activity. The acetylation and phosphorylation of PFK were correlated with shear force and negatively associated with PFK activity. The acetylation and phosphorylation of PK were significantly correlated with each other but showed insignificant correlations with PK activity. Briefly, the phosphorylation and acetylation of HK, PFK and PK coregulate glycolysis through different crosstalk patterns on their activity and this might affect meat quality.

Effects of endurance training on metabolic enzyme activity and transporter protein levels in the skeletal muscles of orchiectomized mice

This study investigated whether endurance training attenuates orchiectomy (ORX)-induced metabolic alterations. At 7 days of recovery after sham operation or ORX surgery, the mice were randomized to remain sedentary or undergo 5 weeks of treadmill running training (15–20 m/min, 60 min, 5 days/week). ORX decreased glycogen concentration in the gastrocnemius muscle, enhanced phosphofructokinase activity in the plantaris muscle, and decreased lactate dehydrogenase activity in the plantaris and soleus muscles. Mitochondrial enzyme activities and protein content in the plantaris and soleus muscles were also decreased after ORX, but preserved, in part, by endurance training. In the treadmill running test (15 m/min, 60 min) after 4 weeks of training, orchiectomized sedentary mice showed impaired exercise performance, which was restored by endurance training. Thus, endurance training could be a potential therapeutic strategy to prevent the hypoandrogenism-induced decline in muscle mitochondrial content and physical performance.

Lactate receptor HCAR1 regulates cell growth, metastasis and maintenance of cancer‑specific energy metabolism in breast cancer cells.

Under aerobic conditions, the preferential use of anaerobic glycolysis by tumour cells leads to a high level of lactate accumulation in tumour microenvironment. Lactate acts not only as a cellular energy source but also as a signalling molecule that regulates cancer cell growth, metastasis and metabolism. It has been reported that a G-protein-coupled receptor for lactate named hydroxycarboxylic acid receptor 1 (HCAR1) is highly expressed in numerous types of cancer, but the detailed mechanism remains unclear. In the present study, it was reported that HCAR1 is highly expressed in breast cancer cells. Genetic deletion of HCAR1 in MCF7 cells leads to reduced cell proliferation and migration. Moreover, it was observed that knockout (KO) of HCAR1 attenuated the expression and activity of phosphofructokinase and hexokinase, key rate-limiting enzymes in glycolysis. Using an extracellular flux analyzer, it was showed that KO of HCAR1 promoted a metabolic shift towards a decreased glycolysis state, as evidenced by a decreased extracellular acidification rate and increased oxygen consumption rate in MCF7 cells. Taken together, our results suggested that lactate acts through HCAR1 as a metabolic regulator in breast cancer cells that may be therapeutically exploited.

Impacts of microplastics and carbamazepine on the shell formation of thick-shell mussels and the underlying mechanisms of action

Forming calcareous exoskeletons is crucial for the health and survival of calcifiers such as bivalves. However, the impacts of waterborne emergent pollutants on this important process remain largely unknown. In this study, the effects of two types of emergent pollutants, microplastics (MPs) and carbamazepine (CBZ), which are ubiquitously present in ocean environments, on shell formation were assessed in the thick-shell mussel (Mytilus coruscus) with a shell regeneration experiment. In addition, their impacts on the in vivo contents of ATP, Ca2+, carbonic anhydrase (CA), and bone morphogenetic protein receptor type-2 (BMPR2), the activity of phosphofructokinase (PFK) and Ca2+-ATPase, and the expression of shell-formation related genes were analyzed. The data collected demonstrated that shell regeneration after mechanical injury was significantly arrested by CBZ and/or MPs. Besides, all the physiological and molecular parameters investigated were markedly suppressed by these two pollutants. Furthermore, synergistic impacts on most of the parameters examined were observed between CBZ and MPs. Our results indicate that these two pollutants may disrupt shell formation by constraining the availability of raw materials and energy, inhibiting the formation of the organic shell matrix, and interfering with the regulation of crystallization, which may have far-reaching impacts on the health of marine calcifiers.

Genistein accelerates glucose catabolism via activation the GPER-mediated cAMP/PKA-AMPK signaling pathway in broiler chickens

Genistein, the most abundance of phytoestrogens in soybeans, has beneficial effects in regulating metabolism-related disease; however, there is few available literatures about whether genistein regulates glucose metabolism that in turn affects the lipid accumulation in animals or humans. The current study showed that genistein promoted glucose uptake by enhancing glucose transporter-2 (GLUT2) protein level; and it also increased the activity of phosphofructokinase-1 (PFK) and pyruvate dehydrogenase (PDH), and the mRNA level of succinate dehydrogenase (SDH) both in broiler chickens or hepatocytes. Moreover, genistein obviously increased the p-LKB1 and p-AMPKα protein levels both in vivo and in vitro. Furthermore, the enhancement of genistein on glucose uptake and catabolism were reversed in hepatocytes pre-treated with AMPK inhibitor Compound C, and the increasing of genistein on the p-LKB1 and p-AMPKα protein levels were also reversed in hepatocytes pre-treated with PKA inhibitor H89. Importantly, the results showed that genistein simultaneously increased the estrogen receptor β (ERβ) and G protein-coupled estrogen receptor (GPER) protein levels, but the elevation effect of genistein on cAMP content was completely reversed in hepatocytes pre-treated with GPER antagonist G15, rather than ERβ inhibitor PHTPP. Meanwhile, the increasing of p-LKB1 and p-AMPKα protein levels induced by genistein were also reversed in hepatocytes pre-treated with G15. Collectively, our data demonstrated that genistein improves glucose metabolism via activating the GPER-mediated cAMP/PKA-AMPK signaling pathway. These findings provide theoretical basis for genistein as a promising nutritional supplemental to alleviate metabolism disorders and related diseases in animals or even humans.

Knockout of TIGAR enhances myocardial phosphofructokinase activity and preserves diastolic function in heart failure.

Hypertension is an important risk factor in the pathogenesis of diastolic dysfunction. Growing evidence indicates that glucose metabolism plays an essential role in diastolic dysfunction. TP53-induced glycolysis and apoptosis regulator (TIGAR) has been shown to regulate glucose metabolism and heart failure (HF). In the present study, we investigated the role of TIGAR in diastolic function and cardiac fibrosis during pressure overload (PO)-induced HF. WT mice subjected to transverse aortic constriction (TAC), a commonly used method to induce diastolic dysfunction, exhibited diastolic dysfunction as evidenced by increased E/A ratio and E/E' ratio when compared to its sham controls. This was accompanied by increased cardiac interstitial fibrosis. In contrast, the knockout of TIGAR attenuated PO-induced diastolic dysfunction and interstitial fibrosis. Mechanistically, the levels of glucose transporter Glut-1, Glut-4, and key glycolytic enzyme phosphofructokinase 1 (PFK-1) were significantly elevated in TIGAR KO subjected to TAC as compared to that of WT mice. Knockout of TIGAR significantly increased fructose 2,6-bisphosphate levels and phosphofructokinase activity in mouse hearts. In addition, PO resulted in a significant increase in perivascular fibrosis and endothelial activation in the WT mice, but not in the TIGAR KO mice. Our present study suggests a necessary role of TIGAR-mediated glucose metabolism in PO-induced cardiac fibrosis and diastolic dysfunction.

In Streptococcus thermophilus, Ammonia from Urea Hydrolysis Paradoxically Boosts Acidification and Reveals a New Regulatory Mechanism of Glycolysis.

ABSTRACTStreptococcus thermophilus is widely used in the dairy industry for the manufacturing of fermented milk and cheeses and probiotic formulations. S. thermophilus evolved from closely phylogenetically related pathogenic streptococci through loss-of-function events counterbalanced by the acquisition of relevant traits, such as lactose and urea utilization for the adaptation to the milk environment. In the context of regressive evolution, the urease gene cluster accounts for 0.9% of the total coding sequence belonging to known functional categories. The fate of ammonia and carbon dioxide derived by urea hydrolysis in several biosynthetic pathways have been depicted, and the positive effect of urease activity on S. thermophilus growth fitness and lactic acid fermentation in milk has been already addressed by several authors. However, the mechanistic effect of urea hydrolysis on the energetic metabolisms of S. thermophilus is still unclear. This study aimed to assess the effect of urease activity on the growth and energy metabolism of Streptococcus thermophilus in milk. In milk, 13C-urea was completely hydrolyzed in the first 150 min of S. thermophilus growth, and urea hydrolysis was accompanied by an increase in cell density and a reduction in the generation time. By using energetically discharged cells with gene transcription and translation blocked, we showed that in the presence of fermentable carbon sources, urease activity, specifically the production of ammonia, could dramatically boost glycolysis and, in cascade, homolactic fermentation. Furthermore, we showed that ammonia, specifically ammonium ions, were potent effectors of phosphofructokinase, a key glycolytic enzyme.IMPORTANCE Finding that ammonia-generating enzymes, such as urease, and exogenous ammonia act on phosphofructokinase activity shed new light on the regulatory mechanisms that govern glycolysis. Phosphofructokinase is the key enzyme known to exert a regulatory role on glycolytic flux and, therefore, ammonia as an effector of phosphofructokinase acts, in cascade, modulating the glycolytic pathway. Apart from S. thermophilus, due to the high conservation of glycolytic enzymes in all branches of the tree of life and being aware of the role of ammonia as an effector of phosphofructokinase, we propose to reevaluate the physiological role of the ammonia production pathways in all organisms whose energy metabolism is supported by glycolysis.

Dietary calcium pyruvate could improve growth performance and reduce excessive lipid deposition in juvenile golden pompano (Trachinotus ovatus) fed a high fat diet.

Excessive lipid deposition in farmed fish is a challenge in the aquaculture industry. To study the effect of dietary calcium pyruvate (CaP) on lipid accumulation in fish, we used a high fat diet (HFD) to establish a lipid accumulation model in juvenile golden pompano (Trachinotus ovatus) and supplemented with 0%, 0.25%, 0.50%, 0.75% and 1.0% CaP (diets D0-D4, respectively). After 8-week feeding in floating cages, dietary CaP significantly improved growth performance, which peaked in fish fed diet D3. Supplementation of CaP significantly decreased whole body lipid content in fish fed D2-D4 and hepatosomatic index and liver lipid content in fish fed D3 and D4. Serum and hepatic antioxidant indices, including glutathione, catalase and superoxide dismutase, showed generally increasing trends in fish fed diets with CaP. In addition, increasing dietary CaP increasingly reduced hepatic activities of hexokinase, phosphofructokinase and pyruvate kinase involved in glycolysis, and increased glycogen contents of the liver and muscle. Dietary CaP up-regulated the liver mRNA expression of pparα, cpt1, hsl and fabp1, but down-regulated expression of srebp-1, fas and acc. In conclusion, 0.75% CaP improved growth performance and reduced excessive lipid deposition by affecting fatty acid synthesis and lipolysis in juvenile T. ovatus fed HFD.

The activation of PPAR\u03b3 enhances Treg responses through up-regulating CD36/CPT1-mediated fatty acid oxidation and subsequent N-glycan branching of T\u03b2RII/IL-2R\u03b1

BackgroundPeroxisome proliferator-activated receptor gamma (PPARγ) is an enhancer of Treg responses, but the mechanisms remain elusive. This study aimed to solve this problem in view of cellular metabolism.MethodsThree recognized PPARγ agonists (synthetic agonist: rosiglitazone; endogenous ligand: 15d-PGJ2; natural product: morin) were used as the tools to activate PPARγ. The fatty acid oxidation (FAO) was evaluated through the detection of fatty acid uptake, oxygen consumption rate, mitochondrial mass, mitochondrial membrane potential and acetyl-CoA level. The involvement of UDP-GlcNAc/N-linked glycosylation axis and the exact role of PPARγ in the action of PPARγ agonists were determined by flow cytometry, Q-PCR, western blotting, a commercial kit for enzyme activity and CRISPR/Cas9-mediated knockout.ResultsRosiglitazone, 15d-PGJ2 and morin all increased the frequency of CD4+Foxp3+ Treg cells generated from naïve CD4+ T cells, boosted the transcription of Foxp3, IL-10, CTLA4 and TIGIT, and facilitated the function of Treg cells. They significantly promoted FAO in differentiating Treg cells by up-regulating the levels of CD36 and CPT1 but not other enzymes involved in FAO such as ACADL, ACADM, HADHA or HADHB, and siCD36 or siCPT1 dampened PPARγ agonists-promoted Treg responses. Moreover, PPARγ agonists enhanced UDP-GlcNAc biosynthesis and subsequent N-linked glycosylation, but did not affect the expressions of N-glycan branching enzymes Mgat1, 2, 4 and 5. Notably, the enzyme activity of phosphofructokinase (PFK) was inhibited by PPARγ agonists and the effect was limited by siCD36 or siCPT1, implying PFK to be a link between PPARγ agonists-promoted FAO and UDP-GlcNAc biosynthesis aside from acetyl-CoA. Furthermore, PPARγ agonists facilitated the cell surface abundance of TβRII and IL-2Rα via N-linked glycosylation, thereby activating TGF-β/Smads and IL-2/STAT5 signaling, and the connection between N-linked glycosylation and Treg responses was revealed by tunicamycin. However, the increased surface abundance of CD36 was demonstrated to be mainly owing to PPARγ agonists-up-regulated overall expression. Finally, PPARγ antagonist GW9662 or CRISPR/Cas9-mediated knockout of PPARγ constrained the effects of rosiglitazone, 15d-PGJ2 and morin, confirming the exact role of PPARγ.ConclusionsThe activation of PPARγ enhances Treg responses through up-regulating CD36/CPT1-mediated fatty acid oxidation and subsequent N-glycan branching of TβRII/IL-2Rα, which is beneficial for inflammatory and autoimmune diseases.Graphical abstract3dQZ3YAwPWWgRqCN5uuEUmVideo

A novel description of the Vastus lateralis morphology of the Temminck's ground pangolin (Manis temminckii).

The pangolin (Manidae family) is the world's most trafficked animal, yet very little is known about its physiology and metabolism primarily due to its inconspicuous and solitary nature. Skeletal muscle samples from the Vastus lateralis were collected postmortem from a single female Temminck's ground pangolin (Manis temminckii). Samples were analyzed for fiber type composition, fiber size and myosin heavy chain isoform content. The oxidative and glycolytic metabolic capacity was determined through citrate synthase, 3-hydroxyacetyl co A dehydrogenase, creatine kinase, lactate dehydrogenase, phosphofructokinase and glycogen phosphorylase enzyme activities. Lastly, antioxidant capacity was determined through superoxide dismutase and catalase enzyme activities, and the total antioxidant capacity. The pangolin metabolic profile was then compared to other endurance and nonendurance mammals, in which data were standardized relative to human endurance athletes in order to provide context. Slow twitch type I fibers, rich in mitochondria were the predominant fiber type within the pangolin indicating a reliance on oxidative derived energy from fats and carbohydrates. This suggests that the pangolin has a high endurance capability when compared to other wild animals and human endurance athletes. This is the first study to investigate the skeletal muscle physiology and metabolism of any pangolin species, in an attempt to further understand this endangered animal and aid with conservation efforts.

High relative humidity enhances chilling tolerance of zucchini fruit by regulating sugar and ethanol metabolisms during cold storage

High relative humidity (RH) cold room (RH: 98 ± 2%) controlled by a ‘dry fog’ system and normal cold room (RH: 74 ± 2%) were used to store zucchini fruit to investigate the regulation of high RH on chilling tolerance and sugar and ethanol metabolisms under cold stress. High RH storage inhibited chilling injury (CI) and maintained higher activities of sucrose synthase-synthesis and sucrose phosphate synthase and contents of glucose, fructose and sucrose in zucchini fruit. Meanwhile, high RH-stored fruit exhibited lower activities of hexokinase, phosphofructokinase, and pyruvate kinase as compared with the control. Besides, the accumulation of acetaldehyde and ethanol under cold stress was decreased by high RH storage together with the inhibition of the increase in activities of alcohol dehydrogenase, pyruvate decarboxylase and expressions of related genes. Taken together, high RH storage enhanced chilling tolerance in zucchini fruit by maintaining high levels of soluble sugars and inhibiting ethanol production.

Dietary chromium could improve growth, antioxidant capacity, chromium accumulation in tissues and expression of genes involved into glucose and lipid metabolism in juvenile mud crab Scylla paramamosain

Chromium (Cr) is an essential micronutrient for all animals, which is also an important component of glucose tolerance factor. The present study was conducted to evaluate the effect of dietary Cr levels on growth, Cr accumulation in tissues, glucose and lipid metabolism and insulin signal pathway of juvenile mud crab. Six isonitrogenous and isolipidic diets were formulated to contain 1.4 (basal diet), 1.7, 2.0, 2.2, 2.6 and 3.7 mg kg−1 Cr, respectively. The results indicated that crabs fed with 2.2 and 2.6 mg kg−1 Cr diets had the highest percent weight gain (PWG) and specific growth rate (SGR) among all treatments. Cr concentration in hepatopancreas significantly increased with increase of dietary Cr levels. Crabs fed with 3.7 mg kg−1 Cr diet exhibited lower antioxidant enzyme activities and higher malondialdehyde (MDA) in hepatopancreas and hemolymph than those fed the other diets. Crabs fed the diets with 2.2 and 2.6 mg kg−1 Cr showed higher activities of hexokinase (HK), phosphofructokinase (PFK) and pyruvate kinase (PK). The expression levels of genes related to insulin signaling pathway, glycolysis and fat synthesis, glycolipid metabolism (such as phosphatidylinositol 3-kinase (pi3k), hexokinase (hk), and fatty acid synthase (fas)) in hepatopancreas were significantly up-regulated. The expression levels of genes related to gluconeogenesis and lipolysis such as fructose-1,6-bisphosphatase (fbp) and carnitine palmitoyltransferase 1 (cpt1) were significantly down-regulated in crabs fed with optimal Cr diets. In conclusion, the optimal dietary Cr requirement was estimated to be 2.34 mg kg−1 for juvenile mud crab based on two slope broken-line regression analysis of PWG against dietary Cr levels, the results of present study indicated that optimal dietary Cr promoted the growth and Cr accumulation in tissues, and modulated insulin signaling pathway to trigger glycolysis and glycogenesis to maintain glucose homeostasis.

No Differences Between 12 Weeks of Block- vs. Traditional-Periodized Training in Performance Adaptations in Trained Cyclists

The purpose of this study was to compare the effects of 12 weeks load-matched block periodization (BP, n = 14), using weekly concentration of high- (HIT), moderate- (MIT), and low- (LIT) intensity training, with traditional periodization (TP, n = 16) using a weekly, cyclic progressive increase in training load of HIT-, MIT-, and LIT-sessions in trained cyclists (peak oxygen uptake: 58 ± 8 ml·kg−1·min−1). Red blood cell volume increased 10 ± 16% (p = 0.029) more in BP compared to TP, while capillaries around type I fibers increased 20 ± 12% (p = 0.002) more in TP compared to BP from Pre to Post12. No other group differences were found in time-trial (TT) performances or muscular-, or hematological adaptations. However, both groups improved 5 and 40-min TT power by 9 ± 9% (p < 0.001) and 8 ± 9% (p < 0.001), maximal aerobic power (Wmax) and power output (PO) at 4 mmol·L−1 blood lactate (W4mmol), by 6 ± 7 (p = 0.001) and 10 ± 12% (p = 0.001), and gross efficiency (GE) in a semi-fatigued state by 0.5 ± 1.1%-points (p = 0.026). In contrast, GE in fresh state and VO2peak were unaltered in both groups. The muscle protein content of β-hydroxyacyl (HAD) increased by 55 ± 58% in TP only, while both TP and BP increased the content of cytochrome c oxidase subunit IV (COXIV) by 72 ± 34%. Muscle enzyme activities of citrate synthase (CS) and phosphofructokinase (PFK) were unaltered. TP increased capillary-to-fiber ratio and capillary around fiber (CAF) type I by 36 ± 15% (p < 0.001) and 17 ± 8% (p = 0.025), respectively, while BP increased capillary density (CD) by 28 ± 24% (p = 0.048) from Pre to Post12. The present study shows no difference in performance between BP and “best practice”-TP of endurance training intensities using a cyclic, progressively increasing training load in trained cyclists. However, hematological and muscle capillary adaptations may differ.