Pyruvate Kinase Research Articles

Novel Insights Into DLAT's Role in Alzheimer's Disease-Related Copper Toxicity Through Microglial Exosome Dynamics.

Alzheimer's disease (AD) is a complex neurodegenerative disorder, with recent research emphasizing the roles of microglia and their secreted extracellular vesicles in AD pathology. However, the involvement of specific molecular pathways contributing to neuronal death in the context of copper toxicity remains largely unexplored. This study investigates the interaction between pyruvate kinase M2 (PKM2) and dihydrolipoamide S-acetyltransferase (DLAT), particularly focusing on copper-induced neuronal death in Alzheimer's disease. Gene expression datasets were analyzed to identify key factors involved in AD-related copper toxicity. The role of DLAT was validated using 5xFAD transgenic mice, while invitro experiments were conducted to assess the impact of microglial exosomes on neuronal PKM2 transfer and DLAT expression. The effects of inhibiting the PKM2 transfer via microglial exosomes on DLAT expression and copper-induced neuronal death were also evaluated. DLAT was identified as a critical factor in the pathology of AD, particularly in copper toxicity. In 5xFAD mice, increased DLAT expression was linked to hippocampal damage and cognitive decline. Invitro, microglial exosomes were shown to facilitate the transfer of PKM2 to neurons, leading to upregulation of DLAT expression and increased copper-induced neuronal death. Inhibition of PKM2 transfer via exosomes resulted in a significant reduction in DLAT expression, mitigating neuronal death and slowing AD progression. This study uncovers a novel pathway involving microglial exosomes and the PKM2-DLAT interaction in copper-induced neuronal death, providing potential therapeutic targets for Alzheimer's disease. Blocking PKM2 transfer could offer new strategies for reducing neuronal damage and slowing disease progression in AD.

Nonylphenol displays immunotoxicity by triggering hemocyte extracellular traps in Manila clam via ROS burst, ERK pathway and glycolysis

Nonylphenol (NP), an endocrine disruptor, has been demonstrated to be a harmful environmental contaminant and toxic to organisms. In this study, to address concerns regarding the immunotoxicity of NP, we treated clam Ruditapes philippinarum hemocytes with NP in vitro and explored the underlying mechanisms of NP-induced extracellular traps (ETs). NP could induce the formation of hemocytes ETs in a dose-dependent manner. Transcriptomics analysis revealed changes of signaling pathway involved in immunity and energy metabolism in hemocytes after NP stimulation. In this process, both reactive oxygen species (ROS) and myeloperoxidase (MPO) were up-regulated. Moreover, mitogen-activated protein kinase (MAPK) signaling pathway was proved to be activated in the formation of NP-induced ETs, manifested as enhanced phosphorylation of extracellular signal-regulated kinase (ERK) but not p38 or c-Jun N-terminal kinase (JNK). In the presence of U0126, an ERK phosphorylation inhibitor, the NP-induced expression of NADPH oxidase enzyme (NOX) was significantly decreased, which further alleviated the ROS production and ultimately limited the release of ETs. NP exposure increased glucose uptake, along with enhanced activities of glycolysis-related enzymes such as hexokinase (HK) and pyruvate kinase (PK). After inhibiting glycolysis by the inhibitor 2-DG, the formation of NP-induced ETs was significantly suppressed. ERK could regulate mTOR signaling and the PI3K/AKT pathway, potentially directing ETs formation by orchestrating the glycolysis through the activation of key transcription factors c-Myc and HIF-1α. Collectively, the results preliminary confirm that the ERK-NOX-ROS axis and glycolysis are involved in NP-induced ETs formation, contributing to the cellular immunotoxicity in clam.

4D-FastDIA-based quantitative proteomics reveals protein biomarkers linked to flavor traits between triploid and diploid oysters (Crassostrea angulata)

In this study, the 4D-FastDIA-based quantitative proteomics strategy was employed to identify protein biomarkers linked to flavor traits in triploid and diploid oysters (Crassostrea angulata). A comprehensive analysis identified a total of 61,947 peptides and 9,703 protein groups across four sample groups, with 3,679 common proteins shared among all groups. Differentially expressed proteins (DEPs) were found to be significantly associated with flavor-related traits, revealing notable differences in amino acids, lipids, and nucleotides between triploid and diploid oysters. Enrichment analysis further elucidated significant pathways involved in metabolic processes, biosynthesis, and immune responses. Among the DEPs, 864 were correlated with flavor quality traits, highlighting key proteins such as S-adenosylmethionine synthase and pyruvate kinase as potential biomarkers for flavor traits. These findings provide valuable insights into the molecular mechanisms underlying flavor differences in oysters and offer potential biomarkers for evaluating and enhancing oyster quality.

The Activation of PKM2 Induces Pyroptosis in Hippocampal Neurons via the NLRP3/Caspase-1/GSDMD Pathway in Neonatal Rats With Hypoxic-Ischemic Brain Injury.

The presence of hypoxic-ischemic brain damage (HIBD) in neonates triggers a strong neuroinflammatory reaction. Pyroptosis, a programmed cell death mechanism associated with inflammation, plays a crucial role in HIBD. Pyruvate kinase M2 (PKM2) plays a significant role in connecting metabolic processes and inflammatory responses, but whether it affects hippocampus pyroptosis in HIBD is unclear. The aim of this study is to elucidate the role of PKM2 in HIBD and to propose a novel therapeutic approach for neonatal ischemic-hypoxic encephalopathy. In this study, we employed neonatal 7-day-old Sprague Dawley rats to establish a model of HIBD using the Rice-Vannucci surgical technique and a hypoxia device. To inhibit the elevation of PKM2, we utilized the PKM2 inhibitor shikonin. The rats were categorized into four groups: Sham, Shikonin, HIBD, and Shikonin+HIBD. Behavioral tests, hematoxylin eosin staining, immunofluorescence staining, ELISA (IL-1β, IL-18), and LDH were conducted in each group to evaluate neurological function, hippocampal damage, the occurrence of neuronal pyroptosis, and the neuroinflammation. Western blot was used to assess the expression levels of PKM2, NLRP3, Caspase-1, Cleaved Caspase-1, GSDMD, GSDMDN, and IL-1β. The expression of PKM2 elevated in hippocampal tissues of the HIBD model and the localization of PKM2 in the hippocampus was activated in neurons instead of microglia during the HIBD. Meanwhile, the inhibition of PKM2 improved the behavioral test scores and the body weight of rats, the neuronal damage in the CA1 region of hippocampal tissue was also attenuated. In addition, inhibiting PKM2 alleviated neuronal pyroptosis by decreasing the expression of PKM2, NLRP3, Caspase-1, Cleaved Caspase-1, GSDMD, GSDMDN. Furthermore, serum levels of LDH and inflammatory factors IL-1β and IL-18 decrease with PKM2 inhibition. Based on these findings, we can conclude that PKM2 plays a crucial role in regulating hippocampal neuronal pyroptosis of HIBD rats via NLRP3/Caspase-1/GSDMD pathway. Therefore, inhibiting PKM2 could be a promising therapeutic strategy for the treatment of neonatal ischemic-hypoxic encephalopathy.

Non-metabolic enzyme function of pyruvate kinase M2 in breast cancer.

Breast cancer (BC) is a prevalent malignant tumor in women, and its incidence has been steadily increasing in recent years. Compared with other types of cancer, it has the highest mortality and morbidity rates in women. So, it is crucial to investigate the underlying mechanisms of BC development and identify specific therapeutic targets. Pyruvate kinase M2 (PKM2), an important metabolic enzyme in glycolysis, has been found to be highly expressed in BC. It can also move to the nucleus and interact with various transcription factors and proteins, including hypoxia-inducible factor-1α (HIF-1α), signal transducer and activator of transcription 3 (STAT3), β-catenin, cellular-myelocytomatosis oncogene (c-Myc), nuclear factor kappa-light-chain enhancer of activated B cells (NF-κB), and mammalian sterile 20-like kinase 1 (MST1). This interaction leads to non-metabolic functions that control the cell cycle, proliferation, apoptosis, migration, invasion, angiogenesis, and tumor microenvironment in BC. This review provides an overview of the latest advancements in understanding the interactions between PKM2 and different transcription factors and proteins that influence the initiation and progression of BC. It also examined how natural drugs and noncoding RNAs affect various biological processes in BC cells through the regulation of the non-metabolic enzyme functions of PKM2. The findings provide valuable insights for improving the prognosis and developing targeted therapies for BC in the coming years.

Improving sugar and respiratory metabolism in pear wounds by postharvest dipping with chitosan and chitooligosaccharide

Chitosan (CTS) and its degradation product, chitooligosaccharide (COS), promote fruit healing by activating phenylpropanoid metabolism. This study investigates their effects on sucrose metabolism in pear wounds. CTS and COS were found to activate neutral invertase, acid invertase, sucrose synthase, and sucrose phosphate synthase, increasing sucrose, glucose, and fructose levels in fruit wounds. They also enhanced sorbitol dehydrogenase activity and promoted sorbitol accumulation. In addition, CTS and COS improved the activities of hexokinase, phosphofructokinase, and pyruvate kinase, increasing phosphoenolpyruvate and ATP production. They activated glucose-6-phosphate dehydrogenase and increased erythrose-4-phosphate, NADPH, and shikimic acid levels. In conclusion, CTS and COS support the formation of the healing closing layer by supplying carbon skeletons, energy, and reducing power through the activation of sugar and respiratory metabolism during the healing process. Compared to CTS, COS was superior in activating the above metabolisms, which is expected to be widely used as a chitin product in postharvest fruit and vegetable preservation and provide new insights into preserving pear freshness.

Polyphyllin II inhibits breast cancer cell proliferation via the PI3K/Akt signaling pathway.

Paridis Rhizoma saponins (PRS) are significant components of Rhizoma Paridis and have inhibitory effects on various tumors, such as bladder, breast, liver and colon cancer. Polyphyllin II (PPII), one of the PRS, has an unclear effect on breast cancer. The present study aimed to explore the effect and mechanism of PPII in breast cancer. A network pharmacology approach was employed to predict the core components and breast cancer‑related targets of PRS. Moreover, a xenograft tumor model was established to determine the anti‑breast cancer effect of PPII in vivo. The viability of MDA‑MB‑231 cells was determined by a Cell Counting Kit‑8 assay. Apoptosis was analyzed using annexin V/PI double staining. Additionally, Transwell and scratch assays were performed to evaluate invasion and migration. The potential mechanism was predicted by Kyoto Encyclopedia of Genes and Genomes enrichment analysis and molecular docking analysis and verified by western blot analysis. The effect of PPII on aerobic glycolysis in breast cancer cells was detected by lactic acid and pyruvate kits and Western blotting of glycolytic rate‑limiting enzymes. Network pharmacology analysis revealed 26 core targets involved in breast cancer and that PPII was the core active component of PRS. The in vivo studies showed that PPII could inhibit the growth of breast cancer in mice. In vitro experiments confirmed that PPII induced cancer cell apoptosis and inhibited invasion and migration. Furthermore, PPII was capable of suppressing the expression of key proteins in the PI3K/Akt signaling pathway, reducing the generation of aerobic glycolytic products, and diminishing the protein expression levels of hexokinase 2 and pyruvate kinase M2. The results indicated that PPII inhibited aerobic glycolysis in breast cancer cells through the PI3K/Akt signaling pathway, thereby inhibiting breast cancer growth.

Fructose shields human colorectal cancer cells from hypoxia-induced necroptosis

Recent studies have shown that high dietary fructose intake enhances intestinal tumor growth in mice. Our previous work indicated that glucose enables hypoxic colorectal cancer (CRC) cells to resist receptor-interacting protein (RIP)-dependent necroptosis. Despite having the same chemical formula, glucose and fructose are absorbed through different transporters yet both can enter the glycolytic metabolic pathway. The excessive intake of dietary fructose, leading to its overflow into the colon, allows colonic cells to absorb fructose apically. This study explores the mechanisms behind apical fructose-mediated death resistance in CRC cells under hypoxic stress. Utilizing three CRC cell lines (Caco-2, HT29, and T84) under normoxic and hypoxic conditions with varying fructose concentrations, we assessed lactate dehydrogenase (LDH) activity, RIP1/3 complex formation (a necroptosis marker), and cell integrity. We investigated the role of fructose in glycolytic-mediated death resistance using glycolytic inhibitors iodoacetate (IA, a glycolytic inhibitor to glyceraldehyde 3-phosphate dehydrogenase), and UK5099 (UK, an inhibitor to mitochondrial pyruvate carrier). Our findings reveal that apical fructose prevents the hypoxia-induced RIP-dependent necroptosis in Caco-2 and HT29 cells. Fructose exposure under hypoxia also preserved epithelial integrity. IA, but not UK, blocked fructose-mediated glycolytic metabolite production and necrosis, indicating that anaerobic glycolytic metabolites facilitate death resistance. Notably, fructose treatment upregulated pyruvate kinase (PK)-M1 mRNA in hypoxic Caco-2 and HT29 cells, while PKM2 upregulation was exclusive to HT29 cells. In conclusion, apical fructose utilization through glycolysis effectively inhibits hypoxia-induced RIP-dependent necroptosis in CRC cells, shedding light on potential metabolic adaptation mechanisms in the tumor microenvironment and suggesting novel targets for therapeutic intervention.

Pde3a and Pde3b regulation of murine pulmonary artery smooth muscle cell growth and metabolism.

A role for metabolically active adipose tissue in pulmonary hypertension (PH) pathogenesis is emerging. Alterations in cellular metabolism in metabolic syndrome are triggers of PH-related vascular dysfunction. Metabolic reprogramming in proliferative pulmonary vascular cells causes a metabolic switch from oxidative phosphorylation to glycolysis. PDE3A and PDE3B subtypes in the regulation of metabolism in pulmonary artery smooth muscle cells (PASMC) are poorly understood. We previously found that PDE3A modulates the cellular energy sensor, AMPK, in human PASMC. We demonstrate that global Pde3a knockout mice have right ventricular (RV) hypertrophy, elevated RV systolic pressures, and metabolic dysfunction with elevated serum free fatty acids (FFA). Therefore, we sought to delineate Pde3a/Pde3b regulation of metabolic pathways in PASMC. We found that PASMC Pde3a deficiency, and to a lesser extent Pde3b deficiency, downregulates AMPK, CREB and PPARγ, and upregulates pyruvate kinase dehydrogenase expression, suggesting decreased oxidative phosphorylation. Interestingly, siRNA Pde3a knockdown in adipocytes led to elevated FFA secretion. Furthermore, PASMC exposed to siPDE3A-transfected adipocyte media led to decreased α-SMA, AMPK and CREB phosphorylation, and greater viable cell numbers compared to controls under the same conditions. These data demonstrate that deficiencies of Pde3a and Pde3b alter pathways that affect cell growth and metabolism in PASMC.

Aqueous Extract of Rhubarb Promotes Hepatotoxicity via Facilitating PKM2-Mediated Aerobic Glycolysis in a Rat Model of Diethylnitrosamine-Induced Liver Cancer.

To identify the polar parts in Rhubarb that cause hepatotoxicity and explore the underlying mechanisms. The rat model of liver cancer was established by gavage of diethylnitrosamine (DEN; 0.002 g/rat) for 14weeks. Starting from the 11th week, Rhubarb granule (4 g/kg), aqueous, ethyl acetate and n-butanol extract of Rhubarb or Rhein equivalent to a dose of 4 g/kg Rhubarb granule were administered intragastrically for 4 consecutive weeks. Liver tissues from rats treated with DEN and Rhubarb granules were used for non-targeted metabolomics analysis. The correlation between pyruvate kinase isozyme type M2 (PKM2) expression level and the progress and prognosis of hepatocellular carcinoma (HCC) was evaluated through bioinformatics analysis based on TCGA database. Liver tissues and blood samples from rats treated with DEN and aqueous, ethyl acetate and n-butanol extract of Rhubarb were used for the screening of hepatotoxic polar parts of Rhubarb. The liver injuries were evaluated by the changes in pathology, liver function, and the expression levels of proliferating cell nuclear antigen (PCNA) and transforming growth factor beta1 (TGF-β1). The mechanism studies focus on PKM2 expression, and the metabolic reprogramming via detecting the activities of lactate dehydrogenase A (LDHA) and isocitrate dehydrogenase (ICDH). Furthermore, molecular docking analysis was performed to validate the target interaction between Rhein and PKM2, and the hepatotoxicity of Rhein was evaluated by testing liver function in the DEN-induced liver cancer model. The non-targeted metabolomics analysis revealed that Rhubarb promoted aerobic glycolysis in the rat model of DEN-induced liver cancer. And bioinformatics analysis revealed that high PKM2 expression was closely related to the progression and poor prognosis of HCC. In vivo studies indicated that the aqueous extract of Rhubarb, but not ethyl acetate and n-butanol extract, promoted the liver injuries induced by DEN. The mechanism study showed that the aqueous extract of Rhubarb increased the expression of PKM2 and promoted aerobic glycolysis. Moreover, Rhein had a strong binding affinity for PKM2 and aggravated liver injury in the DEN-induced liver cancer model. Aqueous extract of Rhubarb promoted hepatotoxicity via facilitating PKM2-mediated aerobic glycolysis in the rat model of DEN-induced liver cancer.

Pterostilbene Induces Pyroptosis in Breast Cancer Cells through Pyruvate Kinase 2/Caspase-8/Gasdermin C Signaling Pathway.

The incidence and mortality of breast cancer increase year by year, and it is urgent to find high-efficiency and low-toxicity anti-cancer drugs. Pterostilbene (PTE) is a natural product with antitumor activity, but the specific antitumor mechanism is not very clear. Aerobic glycolysis is the main energy supply for cancer cells. Pyroptosis is an inflammatory, programmed cell death. The aim of this study was to investigate the effect of PTE on glycolysis and pyroptosis in EMT6 and 4T1 cells and the specific mechanism, and to elucidate the role of pyruvate kinase 2 (PKM2), a key enzyme in glycolysis, in the antitumor role of PTE. Our study suggested that PTE induced pyroptosis by inhibiting tumor glycolysis. PKM2 played an important role in both the inhibition of glycolysis and the induction of pyroptosis by PTE.

Targeting insulin resistance: myricetin and isorhamnetin from Hardwickia binata, and luteolin from Hedysarum alpinum enhance glucose uptake and AMPK signaling in HepG2 cells

This study examined five plants (Xylopia aethiopica, Agave sisalana, Hardwickia binata, Hedysarum alpinum, and Toxicodendron vernicifluum) for their potential to address insulin resistance in type 2 diabetes. In-vitro assays showed that H. binata leaves and H. alpinum flowers inhibited α-glucosidase and α-amylase while enhancing glucose uptake in normal and insulin-resistant HepG2 cells. Phytochemical screening and SPE purification identified the key constituents responsible for the effects. The chromatographic and spectral analysis confirmed flavonoids in H. binata (myricetin, isorhamnetin, quercetin, kaempferol, and catechin) and H. alpinum (luteolin, quercetin, kaempferol, and apigenin). Myricetin, isorhamnetin, and luteolin significantly increased glucose uptake, enhanced hexokinase and pyruvate kinase activities, and promoted IRec and IRS-1 phosphorylation, modulating insulin signalling. They activated AMPK and Akt, with molecular docking confirming strong AMPK binding. These findings suggest that H. binata, H. alpinum, and their flavonoids are promising candidates for managing insulin resistance and type 2 diabetes, warranting further research.

The Role of Pyruvate Kinase M2 Posttranslational Modification in the Occurrence and Development of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is one of the deadly malignant tumors that directly leads to the death of nearly one million people worldwide every year, causing a serious burden on society. In the presence of sufficient oxygen, HCC cells rapidly generate energy through aerobic glycolysis, which promotes tumor cell proliferation, immune evasion, metastasis, angiogenesis, and drug resistance. Pyruvate kinase M2 (PKM2) is a key rate-limiting enzyme in glycolysis. In recent years, studies have found that PKM2 not only exerts pyruvate kinase activity in the process of glucose metabolism, but also exerts protein kinase activity in non-metabolic pathways to affect tumor cell processes, and its activity is flexibly regulated by various posttranslational modifications such as acetylation, phosphorylation, lactylation, ubiquitination, SUMOylation, and so forth. This review summarizes the role of posttranslational modifications of PKM2-related sites in the development of HCC.

Deficiency of metabolic regulator PKM2 activates the pentose phosphate pathway and generates TCF1+ progenitor CD8+ T cells to improve immunotherapy.

TCF1high progenitor CD8+ T cells mediate the efficacy of immunotherapy; however, the mechanisms that govern their generation and maintenance are poorly understood. Here, we show that targeting glycolysis through deletion of pyruvate kinase muscle 2 (PKM2) results in elevated pentose phosphate pathway (PPP) activity, leading to enrichment of a TCF1high progenitor-exhausted-like phenotype and increased responsiveness to PD-1 blockade in vivo. PKM2KO CD8+ T cells showed reduced glycolytic flux, accumulation of glycolytic intermediates and PPP metabolites and increased PPP cycling as determined by 1,2-13C glucose carbon tracing. Small molecule agonism of the PPP without acute glycolytic impairment skewed CD8+ T cells toward a TCF1high population, generated a unique transcriptional landscape and adoptive transfer of agonist-treated CD8+ T cells enhanced tumor control in mice in combination with PD-1 blockade and promoted tumor killing in patient-derived tumor organoids. Our study demonstrates a new metabolic reprogramming that contributes to a progenitor-like T cell state promoting immunotherapy efficacy.

Polysaccharides from Tremella Fuciformis Enhance Glucose and Lipid Metabolism in HepG2 Cells Through Activating the AMPK Signaling Pathway.

Polysaccharides have gained substantial attention for their diverse biological activities. The present study was conucted to elucidate the effects and molecular mechanisms of Tremella fuciformis-derived polysaccharides (PTP-3a) on glucose and lipid metabolism in palmitic acid (PA) - treated HepG2 cells. Multiple parameters were assessed following PTP-3a treatment, including lipid accumulation, glycogen content, glucose consumption, and enzyme activities, including pyruvate kinase (PK) and hexokinase (HK). Additionally, the expression levels of genes associated with glucose and lipid metabolism was evaluated using western blot analysis. PTP-3a effectively inhibited lipid accumulation, promoted the glucose consumption, increased the amount of cellular glycogen, and enhanced PK and HK activities in PA-treated cells. Furthermore, PTP-3a induced a significant increase in the p-AMPK/AMPK ratio and the expression level of PPARa, while decreasing the expression levels of SREBP, FAS, ACC, and SOCS3. In conclusion, these findings suggested that PTP-3a exerted beneficial effects on glucose and lipid metabolism by activating the AMPK signaling pathway, resulting in the inhibition of lipogenesis, promotion of fatty acid oxidation, and enhancement of cellular glycogen synthesis and glycolysis. These findings hold clinical relevance and provide a foundation for potential treatments for non-alcoholic fatty liver disease (NAFLD) and and related metabolic disorders.

Efficient nata de coco production of Komagataeibacter nataicola driven by microbiota-fermented coconut water: Biological and structural characteristics

Nata de coco is a traditional fermented food, chemically known as bacterial cellulose (BC), widely used in the food industry, personal care, and biomaterial areas. In the nata de coco industry, microbiota-fermented coconut water (FCW) has been regarded as a highly efficient medium for improving BC synthesis by Komagataeibacter spp. However, its effects on the biological properties of the strains of Komagataeibacter and structural characteristics of BC remain unknown, resulting in unstable production and BC quality. This study aimed to explore the biological properties of K. nataicola, the characteristics of BC structure and reasons for the efficiency under the effect of FCW. The results showed that FCW could increase the BC yield of K. nataicola by 2.03–4.75 times, the uridine diphosphate glucose pyrophosphorylase activity by 10–45.3 times, and the hexokinase, phosphofructokinase, or pyruvate kinase activities by 2 times. The extracellular metabolite profile of K. nataicola showed that its glycolysis/gluconeogenesis pathway was significantly stimulated and the utilization of erythritol obviously increased. These phenomena might be attributed to the use of ethanol, lactate, xylitoland mannitol in FCW by K. nataicola. In addition, the structural characteristics of BC produced in FCW included higher number of hydrogen bonds, thinner fibrils, a looser three-dimensional network, improved porosity and greater mechanical strength. This study offered novel insights into the BC synthesis of K. nataicola and the BC structural characteristics cultivated in FCW, providing valuable references for the efficient production and potential applications of both nata de coco and BC.

Improving hydrolysis and acidification by regulating the oxygen status and solid content of lignocellulosic waste: Emphasis on the unsealed environment and harnessing the potential of microbial communities

The recalcitrant nature of lignocellulosic raw materials poses a challenge for current biogas plant operations, where hydrolysis and acidification (HA) are the rate-determining steps. To explore the HA balance and microbial mechanisms according to oxygen status and in conditions with relatively high solid content, we made a simple change to build an unsealed reactor. We found that the hemicellulose degradation rate increased by 149.3 % under unsealed conditions. Under microaerobic conditions (Mi), HA with more than 10 % total solid (TS) content exhibited a carbon loss rate of < 10 % in the first 7 days. In the 15 % TS Mi treatment, the organic acid conversion coefficient was 0.04, and the concentration increased to 7.4 g/L within 2 days. A high solid content was the key factor for ensuring efficient organic acid production in the early stage, whereas Mi affected the middle stage. Multiple methods revealed that abundant Prevotella and Clostridium, as well as rare Bifidobacterium, Sporacetigenium, and specific bacteria, had substantial effects on efficient organic acid production and HA balance. The correlation with Mi was 0.57–0.83. The abundance of hemicellulase genes increased by 18.41 %, and the abundance of pyruvate kinase in Mi increased by 15.55 % compared with the sealed condition, demonstrating that Mi can provide complementary advantages in the HA process. The findings in this study improved the operational quality of conventional HA and anaerobic digestion.

Influence of Starvation on Biochemical, Physiological, Morphological, and Transcriptional Responses Associated with Glucose and Lipid Metabolism in the Liver of Javelin Goby (Synechogobius hasta).

In this study, the influence of fasting on hepatic glucose and lipid metabolism was explored by examining biochemical, antioxidative, and morphological indicators and transcriptional expression in the liver of javelin goby (Synechogobius hasta) after 0, 3, 7, or 14 days of starvation. Marked reductions in hepatic glycogen and triglycerides occurred from the seventh day of starvation until the end of the trial (p < 0.05). However, no alterations in hepatic cholesterol or protein were detected throughout the entire experiment (p > 0.05). During fasting, the activities of pyruvate kinase, lactate dehydrogenase, and glycogen phosphorylase a all rose firstly and then fell (p < 0.05). The activities of hepatic fatty acid synthase and acetyl-CoA carboxylase were minimized to their lowest levels at the end of food deprivation (p < 0.05), while lipase was elevated after 7-14 days of fasting (p < 0.05). Catalase, glutathione, and the total antioxidative capacity were increased and maintained their higher values in the later stage of fasting (p < 0.05), whereas malondialdehyde was not significantly changed (p > 0.05). Hepatic vein congestion, remarkable cytoplasmic vacuoles, and irregular cell shape were present in S. hasta which endured 3-7 days of fasting and were less pronounced when food shortage was prolonged. In terms of genes associated with glucose and lipid metabolism, the hepatic phosphofructokinase gene was constantly up-regulated during fasting (p < 0.05). However, the mRNA levels of glycogen synthase and glucose-6-phosphatase were obviously lower when the food scarcity extended to 7 days or more (p < 0.05). Fatty acid synthase, stearoyl-CoA desaturase 1, and peroxisome proliferator-activated receptor γ were substantially down-regulated in S. hasta livers after 7-14 days of food deprivation (p < 0.05). However, genes involved in lipolysis and fatty acid transport were transcriptionally enhanced to varying extents and peaked at the end of fasting (p < 0.05). Overall, starvation lasting 7 days or more could concurrently mobilize hepatic carbohydrates and fat as energy resources and diminished their hepatic accumulation by suppressing biosynthesis and enhancing catabolism and transport, ultimately metabolically and structurally perturbing the liver in S. hasta. This work presents preliminary data on the dynamic characteristics of hepatic glucose and lipid metabolism in S. hasta in response to starvation, which may shed light on the sophisticated mechanisms of energetic homeostasis in fish facing nutrient unavailability and may benefit the utilization/conservation of S. hasta.

Integration of metabolomic and transcriptomic analyses reveals novel regulatory functions of the ChREBP transcription factor in energy metabolism.

Carbohydrate Response Element-Binding Protein (ChREBP) is a transcription factor that activates key genes involved in glucose, fructose, and lipid metabolism in response to carbohydrate feeding, but its other potential roles in metabolic homeostasis have not been as well studied. We used liver-selective GalNAc-siRNA technology to suppress expression of ChREBP in rats fed a high fat/high sucrose diet and characterized hepatic and systemic responses by integrating transcriptomic and metabolomic analyses. GalNAc-siChREBP-treated rats had lower levels of multiple short-chain acyl CoA metabolites compared to rats treated with GalNAc-siCtrl containing a non-targeting siRNA sequence. These changes were related to a sharp decrease in free CoA levels in GalNAc-siChREBP treated-rats, accompanied by lower expression of transcripts encoding enzymes and transporters involved in CoA biosynthesis. These activities of ChREBP likely contribute to its complex effects on hepatic lipid and energy metabolism. While core enzymes of fatty acid (FA) oxidation are induced by ChREBP knockdown, accumulation of liver acylcarnitines and circulating ketones indicate diversion of acetyl CoA to ketone production rather than complete oxidation in the TCA cycle. Despite strong suppression of pyruvate kinase and activation of pyruvate dehydrogenase, pyruvate levels were maintained, likely via increased expression of pyruvate transporters, and decreased expression of lactate dehydrogenase and alanine transaminase. GalNAc-siChREBP treatment increased hepatic citrate and isocitrate levels while decreasing levels of distal TCA cycle intermediates. The drop in free CoA levels, needed for the 2-ketoglutarate dehydrogenase reaction, as well as a decrease in transcripts encoding the anaplerotic enzymes pyruvate carboxylase, glutamate dehydrogenase, and aspartate transaminase likely contributed to these effects. GalNAc-siChREBP treatment caused striking increases in PRPP and ZMP/AICAR levels, and decreases in GMP, IMP, AMP, NaNM, NAD(P), and NAD(P)H levels, accompanied by reduced expression of enzymes that catalyze late steps in purine and NAD synthesis. ChREBP suppression also increased expression of a set of plasma membrane amino acid transporters, possibly as an attempt to replenish TCA cycle intermediates. In sum, combining transcriptomic and metabolomic analyses has revealed regulatory functions of ChREBP that go well beyond its canonical roles in control of carbohydrate and lipid metabolism to now include mitochondrial metabolism and cellular energy balance.

PKM1 Regulates the Expression of Autophagy and Neuroendocrine Markers in Small Cell Lung Cancer

Small cell lung cancer (SCLC) is known as recalcitrant cancer with high malignancy and heterogeneity. Immunotherapy has changed the treatment pattern of extensive-disease SCLC (ED-SCLC), but the beneficiary population is limited. Therefore, exploring new therapeutic strategies is an urgent clinical problem to be solved for SCLC. SCLC is characterized by highly active glycolytic metabolism and pyruvate kinase M1 (PKM1) is one of the isozymes of PK, an important rate-limiting enzyme in glycolysis pathway. Previous studies have shown that PKM1 is related to autophagy and drug sensitivity, however, how PKM1 regulates drug sensitivity in SCLC and its mechanism remain unclear. The aim of this study was to investigate the biological functions of PKM1 in SCLC, including its effects on proliferation, migration, autophagy, drug sensitivity, and expression of neuroendocrine (NE)-related markers in SCLC. Western blot was used to detect the expression level of PKM1 in SCLC cells. PKM1 gene-overexpressed SCLC cell lines were constructed by stable lentivirus transfection. Proliferation of cells and drug sensitivity were detected by MTT, and migration ability of cells was determined by Transwell. The level of autophagy was detected by flow cytometry. Western blot was used to determine the expression levels of NE-related proteins. PKM1 was differentially expressed among various SCLC cell lines, and was lower in H1092 cells (P<0.01). Compared with the control group, there was no significant difference in proliferation level of PKM1 overexpressing H1092 cell, but the migration ability was significantly increased (P<0.001), the drug sensitivity was reduced, and the level of autophagy was inhibited (P<0.001). Additionally, overexpression of PKM1 could upregulate the expression of non-neuroendocrine (non-NE)-related proteins (P<0.01) and decrease the expression of NE-related proteins (P<0.01). PKM1 was differentially expressed in SCLC cell lines, and high expression of PKM1 did not affect the proliferation, but affected the migration of SCLC cells. PKM1 might affect drug sensitivity by inhibiting autophagy and regulating the expression of NE markers. These results provide a theoretical basis for exploring the role of PKM1 in SCLC.