Expression Levels Of Several Enzymes Research Articles

Abstract 3018: Targeting lipid metabolism and diet in advanced prostate cancer

Abstract Lipid metabolism reprogramming provides the energetic needs of prostate cancer cells (PCa) to support growth and cell progress through the cell cycle, sustaining a highly proliferative phenotype by increasing the expression levels of several enzymes involved in fatty acid (FAs) synthesis. Fatty acid synthase (FASN), a key enzyme in de novo fatty acid lipogenesis (DNL), is upregulated in metastatic, castration-resistant prostate cancer (CRPC), and its overexpression is related to carcinogenesis, growth, and metastasis. We previously demonstrated that FASN inhibitors show antitumor activity suppressing PCa cell proliferation and slowing tumor growth, both in vitro and in vivo, via rewiring of lipid cancer metabolism and inducing endoplasmic reticulum stress.We observed that impairment of DNL increases polyunsaturated fatty acids (PUFA) uptake as a compensatory mechanism of survival. Exogenously acquired PUFAs cause the remodeling of the prostate cancer cell lipidome, manifesting an enhanced acyl chain unsaturation across several lipid species. Lipid peroxidation and ROS accumulation are also observed, suggesting cell susceptible to oxidative damage following FASN blockade. Alterations in metabolic pathways cause a higher uptake of acetate and glutamine in PCa cells, as assessed by 14C-nutrient incorporation following FASN inhibition. Mitochondrial dysfunction is induced by reduction in several respiratory parameters, including ATP production and basal respiration. This may be, at least in part, the result of a reduction in phosphatidylglycerol, a cardiolipin precursor, altering mitochondrial membrane composition. Supplementation with the PUFA docosahexaenoic acid (DHA) increases phosphatidylcholine levels and total phospholipid acyl chain unsaturation, which can lead to membrane disorganization in prostate cancer cells. Superoxide anion production is enhanced by DHA treatment, as expected, and increased levels of glucose and palmitate oxidation by mitochondria are observed. Interestingly, AR and AR-V7 protein expression, as well as c-MYC and PSA, is reduced with DHA treatment. We then evaluated how de novo lipid synthesis inhibition combined with manipulation of the exogenous supply of fatty acids could impair tumor cell progression. Due to the obligate cell dependence on PUFA following DNL suppression, we combined FASN and Acetyl-CoA carboxylase (ACC) inhibitors with DHA supplementation and observed a significant cell growth inhibition in CRPC cell lines and prostate cancer organoids.Altogether, our results suggest a novel mechanism to inhibit the growth of castration-resistant prostate cancer, combining lipogenesis inhibition with PUFA-enriched diet to overcome advanced disease. Citation Format: Silvia D. Rodrigues, Caroline Fidalgo Ribeiro, Guilherme H. Tamarindo, Hubert Pakula, Massimo Loda. Targeting lipid metabolism and diet in advanced prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3018.

Metabolic changes in adipose tissues in response to β3 -adrenergic receptor activation in mice.

Brown and beige adipocytes dissipate energy as heat. Thus, the activation of brown adipocytes and the emergence of beige adipocytes in white adipose tissue (WAT) are suggested to be useful for preventing and treating obesity. Although β3 -adrenergic receptor activation is known to stimulate lipolysis and activation of brown and beige adipocytes, fat depot-dependent changes in metabolite concentrations are not fully elucidated. The current study examined the effect of treatment with CL-316,243, a β3 -adrenergic receptor agonist, on the relative abundance of metabolites in interscapular brown adipose tissue (iBAT), inguinal WAT (ingWAT), and epididymal WAT (epiWAT). Intraperitoneal injection of CL-316,243 (1 mg/kg) for 3 consecutive days increased the relative abundance of several glycolysis-related metabolites in all examined fat depots. The cellular concentrations of metabolites involved in the citric acid cycle and of free amino acids were also increased in epiWAT by CL-316,243. CL-316,243 increased the expression levels of several enzymes and transporters related to glucose metabolism and amino acid catabolism in ingWAT and iBAT but not in epiWAT. CL-316,243 also induced the emergence of more beige adipocytes in ingWAT than in epiWAT. Furthermore, adipocytes surrounded by macrophages were detected in the epiWAT of mice given CL-316,243. The current study reveals the fat depot-dependent modulation of cellular metabolites in CL-316,243-treated mice, presumably resulting from differential regulation of cell metabolism in different cell populations.

Glycogen Synthesis in Glycogenin 1-Deficient Patients: A Role for Glycogenin 2 in Muscle.

Glycogen storage disease (GSD) type XV is a rare disease caused by mutations in the GYG1 gene that codes for the core molecule of muscle glycogen, glycogenin 1. Nonetheless, glycogen is present in muscles of glycogenin 1-deficient patients, suggesting an alternative for glycogen buildup. A likely candidate is glycogenin 2, an isoform expressed in the liver and heart but not in healthy skeletal muscle. We wanted to investigate the formation of glycogen and changes in glycogen metabolism in patients with GSD type XV. Two patients with mutations in the GYG1 gene were investigated for histopathology, ultrastructure, and expression of proteins involved in glycogen synthesis and metabolism. Apart from occurrence of polyglucosan (PG) bodies in few fibers, glycogen appeared normal in most cells, and the concentration was normal in patients with GSD type XV. We found that glycogenin 1 was absent, but glycogenin 2 was present in the patients, whereas the opposite was the case in healthy controls. Electron microscopy revealed that glycogen was present between and not inside myofibrils in type II fibers, compromising the ultrastructure of these fibers, and only type I fibers contained PG bodies. We also found significant changes to the expression levels of several enzymes directly involved in glycogen and glucose metabolism. To our knowledge, this is the first report demonstrating expression of glycogenin 2 in glycogenin 1-deficient patients, suggesting that glycogenin 2 rescues the formation of glycogen in patients with glycogenin 1 deficiency.

Effect of waterborne zinc exposure on lipid deposition and metabolism in hepatopancreas and muscle of grass carp Ctenopharyngodon idella.

The aim of the present study was to explore the effect of waterborne zinc (control, 0.85, 2.20, 3.10mg/l, respectively) exposure on lipid deposition and metabolism in the hepatopancreas and muscle of grass carp Ctenopharyngodon idella. The lipid content, Zn accumulation, and the activities and expression levels of several enzymes involved in lipid metabolism were determined in hepatopancreas and muscle. Waterborne Zn exposure reduced growth performance and increased Zn accumulation in both tested tissues. In hepatopancreas, Zn exposure increased lipid content, the activities of lipogenic enzymes, such as 6PGD, G6PD, ME, ICDH and FAS, as well as the mRNA expression level of G6PD, 6PGD, ICDH, FAS and SREBP-1. But the activity of CPT I and the mRNA expression of HSL, CPT Iα1a, CPT Iα2a and PPARα were down-regulated by Zn exposure. In contrast, in muscle, waterborne Zn exposure decreased lipid deposition, activities of 6GPD, ICDH and ME, as well as the mRNA expression level of G6PD, ICDH, ME, FAS and SREBP-1. However, the activity of CPT I as well as the mRNA expression level of PPARα, HSL, CPT Iα2a, CPT Iα1b and CPT Iβ were up-regulated by Zn exposure. Our results indicate that waterborne Zn increases lipid content by up-regulating lipogenesis and down-regulating lipolysis in hepatopancreas. But, in muscle, waterborne Zn reduces lipid accumulation by up-regulating lipolysis and down-regulating lipogenesis. Differential patterns of lipid deposition, enzymatic activities and genes' expression indicate the tissue-specific regulatory mechanism in fish.

Effect of waterborne copper on lipid metabolism in hepatopancreas and muscle of grass carpCtenopharyngodon idella

This study aims to determine the effect of waterborne copper (Cu) exposure on lipid metabolism in the hepatopancreas and muscle of grass carp Ctenopharyngodon idella. Grass carp was exposed to four levels of waterborne Cu (control, 0.058, 0.150 and 0.199 mg L−1) for 14 days. The lipid content, Cu accumulation, and the activities and expression levels of several enzymes involved in lipid metabolism were determined in hepatopancreas and muscle. Copper (Cu) accumulation was found in both tested tissues and the content increased with the increase in waterborne Cu concentrations. Waterborne Cu exposure reduced the lipid content and growth performance. In hepatopancreas and muscle, the activities and the mRNA expression of lipogenic enzymes (G6PD, 6PGD, ICDH, ME and FAS) decreased with the increase of waterborne Cu levels. Furthermore, the mRNA expression levels of lipolytic enzymes (HSL and ATGL) and transcription factors (SREBP-1 and PPARα) were down-regulated by Cu exposure. The decrease of lipogenesis prevailed over that of lipolysis, which might lead the reduction in lipid contents. Taken together, this study demonstrates that waterborne Cu exposure can perturb the normal processes of lipid metabolism, which serves to improve our understanding of the mechanisms underlying lipid metabolism and deposition during Cu exposure in fish.

Age-related dysregulation in the lipidome compromises the immune response to the severe acute respiratory coronavirus (VIR5P.1140)

Abstract Age-associated changes have long been known to cause dysregulation in antiviral immune responses. The SARS epidemic of 2002-03 was characteristic in this respect that it exhibited an age-associated increase in mortality. We recently reported that an age-related increase in an eicosanoid, prostaglandin D2 (PGD2) contributed to the observed higher mortality in SARS-CoV infected aged compared to young mice. To examine the factors that resulted in elevated levels of PGD2 in aged mouse lungs, we analyzed the gene expression levels of several enzymes in the eicosanoid pathway. Remarkably, secretory phospholipase A2 group 2D (Pla2g2d), which is preferentially expressed in lung CD11c+ cells, was significantly higher in aged compared to young mice. Lipid profiling of aged and young SARS-CoV-infected mice revealed that unlike young mice, aged mice failed to upregulate many pro-inflammatory lipid mediators although both groups upregulated the pro-resolving ones. Further, aged mice lacking PLA2G2D expression (Pla2g2d-/-) mice exhibited enhanced virus clearance, increased lung DC migration to the draining lymph nodes and increased virus specific T cell responses. Infection of aged Pla2g2d-/- mice with SARS-CoV increased survival from 20% to 80-90%, compared to Pla2g2d+/+ mice. Altogether our data identify a novel role for Pla2g2d in the lungs of aged mice in skewing the lipidome to an anti-inflammatory state and thus predisposing the animal to a worse outcome following SARS-CoV infection.

Effects of insulin and its related signaling pathways on lipid metabolism in the yellow catfish Pelteobagrus fulvidraco.

The influence of insulin on hepatic metabolism in fish is not well understood. The present study was therefore conducted to investigate the effects of insulin on lipid metabolism, and the related signaling pathways, in the yellow catfish Pelteobagrus fulvidraco. Hepatic lipid and intracellular triglyceride (TG) content, the activity and expression levels of several enzymes and the mRNA expression of transcription factors (PPARα and PPARγ) involved in lipid metabolism were determined. Troglitazone, GW6471, fenofibrate and wortmannin were used to explore the signaling pathways by which insulin influences lipid metabolism. Insulin tended to increase hepatic lipid accumulation, the activity of lipogenic enzymes (6PGD, G6PD, ME, ICDH and FAS) and mRNA levels of FAS, G6PD, 6PGD, CPT IA and PPARγ, but down-regulated PPARα mRNA level. The insulin-induced effect could be stimulated by the specific PPARγ activator troglitazone or reversed by the PI3 kinase/Akt inhibitor wortmannin, demonstrating that signaling pathways of PPARγ and PI3 kinase/Akt were involved in the insulin-induced alteration of lipid metabolism. The specific PPARα pathway activator fenofibrate reduced insulin-induced TG accumulation, down-regulated the mRNA levels of FAS, G6PD and 6PGD, and up-regulated mRNA levels of CPT IA, PPARα and PPARγ. The specific PPARα pathway inhibitor GW6471 reduced insulin-induced changes in the expression of all the tested genes, indicating that PPARα mediated the insulin-induced changes of lipid metabolism. The present results contribute new knowledge on the regulatory role of insulin in hepatic metabolism in fish.

Aging bone marrow mesenchymal stromal cells have altered membrane glycerophospholipid composition and functionality

Human mesenchymal stem/stromal cells (hMSC) are increasingly used in advanced cellular therapies. The clinical use of hMSCs demands sequential cell expansions. As it is well established that membrane glycerophospholipids (GPL) provide precursors for signaling lipids that modulate cellular functions, we studied the effect of the donor's age and cell doublings on the GPL profile of human bone marrow MSC (hBMSC). The hBMSCs, which were harvested from five young and five old adults, showed clear compositional changes during expansion seen at the level of lipid classes, lipid species, and acyl chains. The ratio of phosphatidylinositol to phosphatidylserine increased toward the late-passage samples. Furthermore, 20:4n-6-containing species of phosphatidylcholine and phosphatidylethanolamine accumulated while the species containing monounsaturated fatty acids (FA) decreased during passaging. Additionally, in the total FA pool of the cells, 20:4n-6 increased, which happened at the expense of n-3 polyunsaturated FAs, especially 22:6n-3. The GPL and FA correlated with the decreased immunosuppressive capacity of hBMSCs during expansion. Our observations were further supported by alterations in the gene expression levels of several enzymes involved in lipid metabolism and immunomodulation. The results show that extensive expansion of hBMSCs harmfully modulates membrane GPLs, affecting lipid signaling and eventually impairing functionality.

The Bphi008a gene interacts with the ethylene pathway and transcriptionally regulates MAPK genes in the response of rice to brown planthopper feeding.

We examined ways in which the Brown planthopper induced008a (Bphi008a; AY256682) gene of rice (Oryza sativa) enhances the plant's resistance to a specialist herbivore, the brown planthopper (BPH; Nilaparvata lugens). Measurement of the expression levels of ethylene synthases and of ethylene emissions showed that BPH feeding rapidly initiated the ethylene signaling pathway and up-regulated Bphi008a transcript levels after 6 to 96 h of feeding. In contrast, blocking ethylene transduction (using 1-methylcyclopropene) reduced Bphi008a transcript levels in wild-type plants fed upon by BPH. In vitro kinase assays showed that Bphi008a can be phosphorylated by rice Mitogen-activated Protein Kinase5 (OsMPK5), and yeast two-hybrid assays demonstrated that the carboxyl-terminal proline-rich region of Bphi008a interacts directly with this kinase. Furthermore, bimolecular fluorescence complementation assays showed that this interaction occurs in the nucleus. Subsequently, we found that Bphi008a up-regulation and down-regulation were accompanied by different changes in transcription levels of OsMPK5, OsMPK12, OsMPK13, and OsMPK17 in transgenic plants. Immunoblot analysis also showed that the OsMPK5 protein level increased in overexpressing plants and decreased in RNA interference plants after BPH feeding. In transgenic lines, changes in the expression levels of several enzymes that are important components of the defenses against the BPH were also observed. Finally, yeast two-hybrid screening results showed that Bphi008a is able to interact with a b-ZIP transcription factor (OsbZIP60) and a RNA polymerase polypeptide (SDRP).

Hepatic protein expression of lean mice and obese diabetic mice treated with peroxisome proliferator-activated receptor activators.

The peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors that modulate lipid and glucose homeostasis. In the clinic, PPARalpha and PPARgamma agonists are used to treat hypertriglyceridemia and insulin resistance of diabetes, respectively. To gain further insight into the molecular mechanisms underlying the therapeutic actions of these drugs, we have by two-dimensional electrophoresis and mass spectrometry performed a comparative analysis of the hepatic protein expression profiles of lean and obese (ob/ob) mice, and obese mice treated with WY14643 (PPARalpha agonist) or rosiglitazone (PPARgamma agonist). We found that livers from obese mice displayed higher levels of enzymes involved in fatty acid oxidation and lipogenesis compared to lean mice and these differences were further amplified by treatment with both PPAR activators. WY14643 normalized the expression levels of several enzymes involved in glycolysis, gluconeogenesis and amino acid metabolism in the obese mice to the levels of lean mice, whereas rosiglitazone partially normalized levels of enzymes involved in amino acid metabolism. In summary, a classical proteomics approach was successfully used to characterize differences at the hepatic proteome level between lean and obese diabetic mice, to map metabolic pathways affected by treatment, and to discriminate between effects caused by treatment with agonists of the closely related PPARalpha and PPARgamma receptors.