bims-celmim Biomed News
on Cellular and mitochondrial metabolism
Issue of 2024‒03‒31
fifteen papers selected by
Marc Segarra Mondejar, University of Cologne

  1. Biomolecules. 2024 Feb 22. pii: 260. [Epub ahead of print]14(3):
      Anti-ageing biology and medicine programmes are a focus of genetics, molecular biology, immunology, endocrinology, nutrition, and therapy. This paper discusses metabolic therapies aimed at prolonging longevity and/or health. Individual components of these effects are postulated to be related to the energy supply by tricarboxylic acid (TCA) cycle intermediates and free radical production processes. This article presents several theories of ageing and clinical descriptions of the top markers of ageing, which define ageing in different categories; additionally, their interactions with age-related changes and diseases related to α-ketoglutarate (AKG) and succinate SC formation and metabolism in pathological states are explained. This review describes convincingly the differences in the mitochondrial characteristics of energy metabolism in animals, with different levels (high and low) of physiological reactivity of functional systems related to the state of different regulatory systems providing oxygen-dependent processes. Much attention is given to the crucial role of AKG and SC in the energy metabolism in cells related to amino acid synthesis, epigenetic regulation, cell stemness, and differentiation, as well as metabolism associated with the development of pathological conditions and, in particular, cancer cells. Another goal was to address the issue of ageing in terms of individual characteristics related to physiological reactivity. This review also demonstrated the role of the Krebs cycle as a key component of cellular energy and ageing, which is closely associated with the development of various age-related pathologies, such as cancer, type 2 diabetes, and cardiovascular or neurodegenerative diseases where the mTOR pathway plays a key role. This article provides postulates of postischaemic phenomena in an ageing organism and demonstrates the dependence of accelerated ageing and age-related pathology on the levels of AKG and SC in studies on different species (roundworm Caenorhabditis elegans, Drosophila, mice, and humans used as models). The findings suggest that this approach may also be useful to show that Krebs cycle metabolites may be involved in age-related abnormalities of the mitochondrial metabolism and may thus induce epigenetic reprogramming that contributes to the senile phenotype and degenerative diseases. The metabolism of these compounds is particularly important when considering ageing mechanisms connected with different levels of initial physiological reactivity and able to initiate individual programmed ageing, depending on the intensity of oxygen consumption, metabolic peculiarities, and behavioural reactions.
    Keywords:  ageing mechanisms; anti-ageing therapy; bioenergetic mechanisms of ageing; individual ageing processes; individual physiological reactivity; tricarboxylic acid cycle intermediates
  2. Cancer Metab. 2024 Mar 26. 12(1): 10
      BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) has been associated with the host dysmetabolism of branched-chain amino acids (BCAAs), however, the implications for the role of BCAA metabolism in PDAC development or progression are not clear. The mitochondrial catabolism of valine, leucine, and isoleucine is a multistep process leading to the production of short-chain R-CoA species. They can be subsequently exported from mitochondria as short-chain carnitines (SC-CARs), utilized in anabolic pathways, or released from the cells.METHODS: We examined the specificities of BCAA catabolism and cellular adaptation strategies to BCAA starvation in PDAC cells in vitro. We used metabolomics and lipidomics to quantify major metabolic changes in response to BCAA withdrawal. Using confocal microscopy and flow cytometry we quantified the fluorescence of BODIPY probe and the level of lipid droplets (LDs). We used BODIPY-conjugated palmitate to evaluate transport of fatty acids (FAs) into mitochondria. Also, we have developed a protocol for quantification of SC-CARs, BCAA-derived metabolites.
    RESULTS: Using metabolic profiling, we found that BCAA starvation leads to massive triglyceride (TG) synthesis and LD accumulation. This was associated with the suppression of activated FA transport into the mitochondrial matrix. The suppression of FA import into mitochondria was rescued with the inhibitor of the acetyl-CoA carboxylase (ACC) and the activator of AMP kinase (AMPK), which both regulate carnitine palmitoyltransferase 1A (CPT1) activation status.
    CONCLUSIONS: Our data suggest that BCAA catabolism is required for the import of long chain carnitines (LC-CARs) into mitochondria, whereas the disruption of this link results in the redirection of activated FAs into TG synthesis and its deposition into LDs. We propose that this mechanism protects cells against mitochondrial overload with LC-CARs and it might be part of the universal reaction to amino acid perturbations during cancer growth, regulating FA handling and storage.
    Keywords:  BCAA metabolism; Fatty acid/Transport; Fluorescence microscopy; Lipid droplets; Lipidomics; Mitochondria; Pancreatic cancer; Triglycerides
  3. Sci Rep. 2024 03 27. 14(1): 7328
      Organ transplantation is a life-saving procedure affecting over 100,000 people on the transplant waitlist. Ischemia reperfusion injury (IRI) is a major challenge in the field as it can cause post-transplantation complications and limit the use of organs from extended criteria donors. Machine perfusion technology has the potential to mitigate IRI; however, it currently fails to achieve its full potential due to a lack of highly sensitive and specific assays to assess organ quality during perfusion. We developed a real-time and non-invasive method of assessing organs during perfusion based on mitochondrial function and injury using resonance Raman spectroscopy. It uses a 441 nm laser and a high-resolution spectrometer to quantify the oxidation state of mitochondrial cytochromes during perfusion. This index of mitochondrial oxidation, or 3RMR, was used to understand differences in mitochondrial recovery of cold ischemic rodent livers during machine perfusion at normothermic temperatures with an acellular versus cellular perfusate. Measurement of the mitochondrial oxidation revealed that there was no difference in 3RMR of fresh livers as a function of normothermic perfusion when comparing acellular versus cellular-based perfusates. However, following 24 h of static cold storage, 3RMR returned to baseline faster with a cellular-based perfusate, yet 3RMR progressively increased during perfusion, indicating injury may develop over time. Thus, this study emphasizes the need for further refinement of a reoxygenation strategy during normothermic machine perfusion that considers cold ischemia durations, gradual recovery/rewarming, and risk of hemolysis.
  4. Anal Methods. 2024 Mar 27.
      Mitochondria are not only the center of energy metabolism but also involved in regulating cellular activities. Quality and quantity control of mitochondria is therefore essential. Mitophagy is a lysosome-dependent process to clear dysfunctional mitochondria, and abnormal mitophagy can cause metabolic disorders. Therefore, it is necessary to monitor the mitophagy in living cells on a real-time basis. Herein, we developed a pH-responsive fluorescent probe MP for the detection of the mitophagy process using real-time tracing colocalization coefficients. Probe MP showed good pH responses with high selectivity and sensitivity in spectral testing. Probe MP is of positive charge, which is beneficial for accumulating into mitochondrial in living cells. Cells exhibited pH-dependent fluorescence when they were treated with different pH media. Importantly, the changes in the colocalization coefficient between probe MP and Lyso Tracker® Deep Red from 0.4 to 0.8 were achieved in a real-time manner during the mitophagy stimulated by CCCP, starvation and rapamycin. Therefore, combined with the parameter of the colocalization coefficient, probe MP is a potential molecular tool for the real-time tracing of mitophagy to further explore the details of mitophagy.
  5. Int J Mol Sci. 2024 Mar 14. pii: 3273. [Epub ahead of print]25(6):
      Many of the biological processes of the cell, from its structure to signal transduction, involve protein-protein interactions. On this basis, our aim was to identify cellular proteins that interact with ERK5, a serine/threonine protein kinase with a key role in tumor genesis and progression and a promising therapeutic target in many tumor types. Using affinity chromatography, immunoprecipitation, and mass spectrometry techniques, we unveiled an interaction between ERK5 and the mitochondrial glutaminase GLS in pancreatic tumor cells. Subsequent co-immunoprecipitation and immunofluorescence studies supported this interaction in breast and lung tumor cells as well. Genetic approaches using RNA interference techniques and CRISPR/Cas9 technology demonstrated that the loss of ERK5 function led to increased protein levels of GLS isoforms (KGA/GAC) and a concomitant increase in their activity in tumor cells. It is well known that the tumor cell reprograms its intermediary metabolism to meet its increased metabolic needs. In this sense, mitochondrial GLS is involved in the first step of glutamine catabolism, one of the main energy sources in the context of cancer. Our data suggest that ERK5 contributes to the regulation of tumor cell energy metabolism via glutaminolysis.
    Keywords:  ERK5; GLS isoforms; cancer therapy; energy metabolism; glutaminase; mitochondria; proteomics
  6. Sci Rep. 2024 03 23. 14(1): 6938
      As the most malignant tumor, the prognosis of pancreatic cancer is not ideal even in the small number of patients who can undergo radical surgery. As a highly heterogeneous tumor, chemotherapy resistance is a major factor leading to decreased efficacy and postoperative recurrence of pancreatic cancer. In this study, nuclear magnetic resonance (NMR)-based metabolomics was applied to identify serum metabolic characteristics of pancreatic ductal adenocarcinoma (PDAC) and screen the potential biomarkers for its diagnosis. Metabolic changes of patients with different CA19-9 levels during postoperative chemotherapy were also monitored and compared to identify the differential metabolites that may affect the efficacy of chemotherapy. Finally, 19 potential serum biomarkers were screened to serve the diagnosis of PDAC, and significant metabolic differences between the two CA19-9 stratifications of PDAC were involved in energy metabolism, lipid metabolism, amino acid metabolism, and citric acid metabolism. Enrichment analysis of metabolic pathways revealed six shared pathways by PDAC and chemotherapy such as alanine, aspartate and glutamate metabolism, arginine biosynthesis, glutamine and glutamate metabolism, citrate cycle, pyruvate metabolism, and glycogolysis/gluconeogeneis. The similarity between the metabolic characteristics of PDAC and the metabolic responses to chemotherapy provided a reference for clinical prediction of benefits of postoperative chemotherapy in PDAC patients.
    Keywords:  Chemotherapy; Metabolomics; Nuclear magnetic resonance; Pancreatic cancer; Serum
  7. Elife. 2024 Mar 27. pii: RP89938. [Epub ahead of print]12
      Membrane contact sites (MCSs) are junctures that perform important roles including coordinating lipid metabolism. Previous studies have indicated that vacuolar fission/fusion processes are coupled with modifications in the membrane lipid composition. However, it has been still unclear whether MCS-mediated lipid metabolism controls the vacuolar morphology. Here, we report that deletion of tricalbins (Tcb1, Tcb2, and Tcb3), tethering proteins at endoplasmic reticulum (ER)-plasma membrane (PM) and ER-Golgi contact sites, alters fusion/fission dynamics and causes vacuolar fragmentation in the yeast Saccharomyces cerevisiae. In addition, we show that the sphingolipid precursor phytosphingosine (PHS) accumulates in tricalbin-deleted cells, triggering the vacuolar division. Detachment of the nucleus-vacuole junction (NVJ), an important contact site between the vacuole and the perinuclear ER, restored vacuolar morphology in both cells subjected to high exogenous PHS and Tcb3-deleted cells, supporting that PHS transport across the NVJ induces vacuole division. Thus, our results suggest that vacuolar morphology is maintained by MCSs through the metabolism of sphingolipids.
    Keywords:  S. cerevisiae; cell biology; membrane contact sites; sphingolipid; tricalbin; vacuolar morphology
  8. Proc Natl Acad Sci U S A. 2024 Apr 02. 121(14): e2217019121
      Mitochondria constantly fuse and divide for mitochondrial inheritance and functions. Here, we identified a distinct type of naturally occurring fission, tail-autotomy fission, wherein a tail-like thin tubule protrudes from the mitochondrial body and disconnects, resembling autotomy. Next, utilizing an optogenetic mitochondria-specific mechanostimulator, we revealed that mechanical tensile force drives tail-autotomy fission. This force-induced fission involves DRP1/MFF and endoplasmic reticulum tubule wrapping. It redistributes mitochondrial DNA, producing mitochondrial fragments with or without mitochondrial DNA for different fates. Moreover, tensile force can decouple outer and inner mitochondrial membranes, pulling out matrix-excluded tubule segments. Subsequent tail-autotomy fission separates the matrix-excluded tubule segments into matrix-excluded mitochondrial-derived vesicles (MDVs) which recruit Parkin and LC3B, indicating the unique role of tail-autotomy fission in segregating only outer membrane components for mitophagy. Sustained force promotes fission and MDV biogenesis more effectively than transient one. Our results uncover a mechanistically and functionally distinct type of fission and unveil the role of tensile forces in modulating fission and MDV biogenesis for quality control, underscoring the heterogeneity of fission and mechanoregulation of mitochondrial dynamics.
    Keywords:  mitochondrial fission; mitochondrial quality control; optogenetics; photoactivatable proteins; tensile force
  9. Mol Metab. 2024 Mar 21. pii: S2212-8778(24)00053-X. [Epub ahead of print] 101922
      OBJECTIVE: Evaluation of mitochondrial oxygen consumption and ATP production is important to investigate pancreatic islet pathophysiology. Most studies use cell lines due to difficulties in measuring primary islet respiration, which requires specific equipment and consumables, is expensive and poorly reproducible. Our aim was to establish a practical method to assess primary islet metabolic fluxes using standard commercial consumables.METHODS: Pancreatic islets were isolated from mice/rats, dispersed with trypsin, and adhered to pre-coated standard Seahorse or Resipher microplates. Oxygen consumption was evaluated using a Seahorse Extracellular Flux Analyzer or a Resipher Real-time Cell Analyzer.
    RESULTS: We provide a detailed protocol with all steps to optimize islet isolation with high yield and functionality. Our method requires a few islets per replicate; both rat and mouse islets present robust basal respiration and proper response to mitochondrial modulators and glucose. The technique was validated by other functional assays, which show these cells present conserved calcium influx and insulin secretion in response to glucose. We also show that our dispersed islets maintain robust basal respiration levels, in addition to maintaining up to 89% viability after five days in dispersed cultures. Furthermore, OCRs can be measured in Seahorse analyzers and in other plate respirometry systems, using standard materials.
    CONCLUSIONS: Overall, we established a practical and robust method to assess islet metabolic fluxes and oxidative phosphorylation, a valuable tool to uncover basic β-cell metabolic mechanisms as well as for translational investigations, such as pharmacological candidate discovery and islet transplantation protocols.
    Keywords:  Mitochondrial respiration; Oxidative phosphorylation; Oxygen consumption; Pancreatic islets
  10. Metabolites. 2024 Mar 19. pii: 171. [Epub ahead of print]14(3):
      Metabolic reprogramming has emerged as a prominent hallmark of cancer, characterized by substantial alterations in nutrient uptake and intracellular metabolic pathways. Consequently, intracellular metabolite concentrations undergo significant changes which can contribute to tumorigenesis through diverse mechanisms. Beyond their classical roles in regulating metabolic pathway flux, metabolites exhibit noncanonical functions that play a crucial role in tumor progression. In this review, we delve into the nonclassical functions of metabolites in the context of tumor progression, with a particular focus on their capacity to modulate gene expression and cell signaling. Furthermore, we discuss the potential exploitation of these nonclassical functions in the enhancement of cancer therapy.
    Keywords:  metabolic reprogramming; metabolites; noncanonical functions of metabolites; tumor progressions
  11. STAR Protoc. 2024 Mar 25. pii: S2666-1667(24)00136-9. [Epub ahead of print]5(2): 102971
      Efficient metabolism, or the means by which cells produce energy resources, is critical for proper effector function. Here, we present a protocol for examining the bioenergetics and mitochondrial fuel utilization of primary murine autoreactive immunocytes using cellular metabolism-modulating drugs. We describe steps for plate calibration, isolation of primary immunocytes, and Seahorse assay plate preparation. We then detail procedures for performing the XF Cell Mito Stress Test followed by bioenergetics calculations and statistics. For complete details on the use and execution of this protocol, please refer to Wilson et al.1.
    Keywords:  Cell-based Assays; Immunology; Metabolism
  12. Science. 2024 Mar 29. 383(6690): 1484-1492
      Cellular purines, particularly adenosine 5'-triphosphate (ATP), fuel many metabolic reactions, but less is known about the direct effects of pyrimidines on cellular metabolism. We found that pyrimidines, but not purines, maintain pyruvate oxidation and the tricarboxylic citric acid (TCA) cycle by regulating pyruvate dehydrogenase (PDH) activity. PDH activity requires sufficient substrates and cofactors, including thiamine pyrophosphate (TPP). Depletion of cellular pyrimidines decreased TPP synthesis, a reaction carried out by TPP kinase 1 (TPK1), which reportedly uses ATP to phosphorylate thiamine (vitamin B1). We found that uridine 5'-triphosphate (UTP) acts as the preferred substrate for TPK1, enabling cellular TPP synthesis, PDH activity, TCA-cycle activity, lipogenesis, and adipocyte differentiation. Thus, UTP is required for vitamin B1 utilization to maintain pyruvate oxidation and lipogenesis.
  13. Nat Cancer. 2024 Mar;5(3): 384-399
      Cellular redox homeostasis is an essential, dynamic process that ensures the balance between reducing and oxidizing reactions within cells and thus has implications across all areas of biology. Changes in levels of reactive oxygen species can disrupt redox homeostasis, leading to oxidative or reductive stress that contributes to the pathogenesis of many malignancies, including cancer. From transformation and tumor initiation to metastatic dissemination, increasing reactive oxygen species in cancer cells can paradoxically promote or suppress the tumorigenic process, depending on the extent of redox stress, its spatiotemporal characteristics and the tumor microenvironment. Here we review how redox regulation influences tumorigenesis, highlighting therapeutic opportunities enabled by redox-related alterations in cancer cells.
  14. Molecules. 2024 Mar 14. pii: 1298. [Epub ahead of print]29(6):
      Glucose metabolism is a crucial biological pathway maintaining the activation of extra- and intracellular signaling pathways involved in the immune response. Immune cell stimulation via various environmental factors results in their activation and metabolic reprogramming to aerobic glycolysis. Different immune cells exhibit cell-type-specific metabolic patterns when performing their biological functions. Numerous published studies have shed more light on the importance of metabolic reprogramming in the immune system. Moreover, this knowledge is crucial for revealing new ways to target inflammatory pathologic states, such as autoimmunity and hyperinflammation. Here, we discuss the role of glycolysis in immune cell activity in physiological and pathological conditions, and the potential use of inhibitors of glycolysis for disease treatment.
    Keywords:  2-deoxy-D-glucose; D-glucose; D-mannose; autoimmunity; glycolysis; glycolysis inhibitors; immune cells; inflammation
  15. Sci Rep. 2024 03 26. 14(1): 7151
      Nutritional and genetic deficiencies in homocysteine (Hcy) metabolism lead to hyperhomocysteinemia (HHcy) and cause endothelial dysfunction, a hallmark of atherosclerosis, which is a major cause of cardiovascular disease (CVD). Impaired autophagy causes the accumulation of damaged proteins and organelles and is associated with CVD. Biochemically, HHcy is characterized by elevated levels of Hcy and its metabolites, Hcy-thiolactone and N-Hcy-protein. However, whether these metabolites can dysregulate mTOR signaling and autophagy in endothelial cells is not known. Here, we examined the influence of Hcy-thiolactone, N-Hcy-protein, and Hcy on autophagy human umbilical vein endothelial cells. We found that treatments with Hcy-thiolactone, N-Hcy-protein, or Hcy significantly downregulated beclin 1 (BECN1), autophagy-related 5 (ATG5), autophagy-related 7 (ATG7), and microtubule-associated protein 1 light chain 3 (LC3) mRNA and protein levels. We also found that these changes were mediated by upregulation by Hcy-thiolactone, N-Hcy-protein, and Hcy of autophagy-targeting microRNA (miR): miR-21, miR-155, miR-216, and miR-320c. The effects of these metabolites on levels of miR targeting autophagy as well as on the levels of BECN1, ATG5, ATG7, and LC3 mRNA and protein were abrogated by treatments with inhibitors of miR-21, miR-155, miR-216, and mir320c. Taken together, our findings show that Hcy metabolites can upregulate miR-21, miR-155, miR-216, and mir320c, which then downregulate autophagy in human endothelial cells, important for vascular homeostasis.
    Keywords:  Autophagy; Endothelial dysfunction; HUVEC; Homocysteine metabolites; microRNA