Science Note: Cellular metabolism

Targeting metabolic heterogeneity in the tumor microenvironment (TME) is critical in cancer research because metabolic diversity affects cancer and immune cell function, promoting tumor progression and immune suppression. In addition to previous findings,  recent studies have identified acetyl-CoA carboxylase and glycosphingolipid synthesis as key mediators of immune evasion in the TME. In this note, we introduce a review of cancer-promoting mechanisms driven by metabolic heterogeneity in the TME, along with recent findings.
 

Point of Interest
- Metabolic heterogeneity in the tumor microenvironment (TME) arises from cellular diversity and affects the metabolic programs of cancer and immune cells, promoting tumor growth, metastasis, and immune evasion.

- Altered nutrients and signals in the TME suppress effector immune cells and favor regulatory immune cells, contributing to immune suppression.

- Targeting metabolic heterogeneity in the TME is a promising strategy to overcome immune suppression and improve the efficacy of cancer immunotherapy.

Point of Interest
- T cells in the solid tumor microenvironment (TME) must catabolize lipids via mitochondrial fatty acid oxidation (FAO) to provide energy during nutrient stress.

- The solid TME activates acetyl-coenzyme A carboxylase (ACC) in T cells, causing  lipid droplet accumulation in tumor-infiltrating T cells, which inhibits FAO.

- Restricting ACC activity rewires T cell metabolism to maintain energy during TME stress.

Point of Interest
- De novo sphingolipid synthesis is critical for immune evasion in KRAS-driven cancers.

- Blocking glycosphingolipid synthesis increases IFNGR1 levels and enhances IFNγ-induced growth arrest and pro-inflammatory signaling.

- Inhibiting glycosphingolipid production increases natural killer and CD8+ T cell activity and synergizes with checkpoint blockade therapy to enhance anti-tumor responses.

Comparison of metabolic pathway in two types of cancer cells

The dependence of OXPHOS and Glycolysis in two types of cancer cells, HeLa and HepG2, were compared based on Lactate production, ATP levels, and OCR values.

Many cancer cells produce ATP through the glycolytic pathway. On the other hand, it has been recently reported that cancer cells whose glycolytic pathway is suppressed survive by shifting their energy metabolism to OXPHOS by enhancing mitochondrial function, and the dependency of metabolic pathways differs depending on cell lines.

＜Evaluation by Lactate production and ATP levels＞

We confirmed the changes in ATP and Lactate production when ATP synthesis by OXPHOS was inhibited by Oligomycin stimulation and by 2-Deoxy-D-glucose (2-DG) in the glycolytic pathway. The results showed that HeLa cells depend on Glycolysis and HepG2 cells depend on OXPHOS to synthesize ATP.

＜Evaluation by Lactate production and ATP levels＞
When OXPHOS was inhibited in HeLa cells, ATP levels remained unchanged (①), and lactate production increased (②). This suggests that even when OXPHOS is inhibited, glycolysis can be further activated. Conversely, when glycolysis is inhibited, ATP levels decrease significantly (③), indicating that energy production depends on glycolysis. On the other hand, when OXPHOS was inhibited in HepG2 cells, lactate production increased (④), indicating that the cells attempt to compensate for energy production by enhancing glycolysis, but ATP levels still decrease (⑤). This means that even with increased glycolysis, ATP production is not sufficiently compensated. Furthermore, ATP levels decrease more when glycolysis is inhibited (⑥), suggesting that energy production in HepG2 cells depends more on OXPHOS than glycolysis.

Products in Use
   - Glycolysis/OXPHOS Assay Kit

＜Evaluation by OCR value＞

Using the same number of cells, we measured the OCR value when cellular oxygen consumption was promoted by stimulating the cells with FCCP, a mitochondrial uncoupling agent. The results showed that HepG2 cells had higher OCR values than HeLa cells, suggesting a greater dependence on OXPHOS, correlating with the results obtained from ATP level and Lactate production.

Products in Use
   - Extracellular OCR Plate Assay Kit

In recent neurodegenerative disease research has shown that lipid metabolism, such as lipid droplets and lipid peroxides, influences neuronal damage. Here are some of the papers that have identified a relationship between lipid metabolism and the neurotoxic environment.

Neuronal toxicity in neurodegenerative diseases is often associated with the accumulation of lipid droplets and lipid peroxides, which are by-products of lipid metabolism. Lipid droplets in neurons and glia serve as storage for lipids, but excessive accumulation can impair cellular function. Lipid peroxides resulting from oxidative stress are particularly damaging, generating reactive oxygen species that lead to cellular dysfunction and cell death. In tauopathies*, lipid peroxides are transferred to glial cells, exacerbating the inflammation and oxidative stress that contribute to the neurotoxic environment.

*Tauopathy is a neurodegenerative disease characterized by abnormal accumulation of tau protein.

Point of Interest
- ACSL1-positive microglia, characterized by the lipid droplet-associated enzyme ACSL1, are abundant in Alzheimer’s patients with the APOE4/4 genotype.

- Fibrillar Aβ induces ACSL1 expression, triglyceride synthesis and lipid droplet accumulation in APOE-dependent microglia.

- Conditioned media from lipid droplet-containing microglia lead to tau phosphorylation and neurotoxicity, linking genetic risk factors for AD to potential therapeutic targets.

Point of Interest
- ROS accumulation in tauopathies leads to toxic peroxidized lipids that affect neurons and glia in brain disorders.

- Glial endogenous tau is essential for lipid droplet formation and protection against toxic neuronal lipids in flies, rats and human cells.

- Flies lacking glial endogenous tau show motor defects and reduced lifespan, which can be alleviated by antioxidants such as N-acetylcysteine amide.

Point of Interest
- Tauopathy fly and human iPSC neurons show increased lipogenesis and impaired lipid turnover within lipid droplets.

- Unsaturated lipid transfer from tauopathy neurons to microglia leads to lipid droplet accumulation, oxidative stress, inflammation and impaired phagocytosis.

- AMPK inhibits neuronal lipogenesis and supports lipophagy, reducing lipid transfer to microglia, whereas AMPK depletion exacerbates lipid accumulation and neuropathology.

Hepatotoxicity test of drug-induced lipidosis using high-content imaging

Propranolol (a sympathetic β-receptor blocker) was added to a human hepatocellular carcinoma cell line (HepG2 cells), and changes in lipid droplets were observed under a fluorescence microscope. The accumulation of lipid droplets was analyzed by measuring the number, area, and fluorescence intensity of lipid droplets from the acquired microscopic images.

＜Lipid droplet imaging data＞

  Nucleus (blue: Hoechst 33342 ): Ex 385 nm, Em 460 nm  
  Lipid droplet (green: Lipi-Green): Ex 475 nm, Em 535nm

HepG2 cells were treated with propranolol 0, 10, or 30 μmol/l, lipid droplets were stained with Lipi-Green and nuclei with Hoechst 33342 and observed using a fluorescence microscope (Ti2-E inverted microscope). The results showed that lipid droplets increased in a propranolol concentration-dependent manner.

Related Products
   - Lipi-Blue/ Green/ Red/ Deep Red
   - Lipid Droplet Assay Kit - Blue / Deep Red

＜Analysis of lipid droplet accumulation relative to drug treatment concentration＞

High Content Analysis (HCA) microscope system
(Nikon Corporation https://www.microscope.healthcare.nikon.com/)

For details of staining and analysis methods, please refer to "APPLICATION NOTE: Hepatotoxicity test of drug-induced lipidosis using high-content imaging" by Nikon Corporation.

From the fluorescence images obtained, the accumulation of lipid droplets per cell was analyzed by measuring cell number from nuclei and area, number, and fluorescence intensity from lipid droplets. The results showed that the number and area of lipid droplets increased in a propranolol concentration-dependent manner, with lipid droplets forming significantly under concentration conditions of 20 μmol/l or higher. The DS-Qi2 camera, which can capture a wide range of cellular areas in a single shot, was used for imaging, and the EDF module of NIS-Elements software, which can acquire focused images of all lipid droplets, was used for analysis, enabling quantitative analysis with highly reliable statistical data. The EDF module of the NIS-Elements software allows for the acquisition of focused images of all lipid droplets.

Recent research on neurodegenerative diseases is revealing that although neurons require glycolysis, excessive neuronal glycolysis causes cognitive decline. Here are some of the papers showing glycolytic activity in neurons and neurodegenerative diseases.

Neurons require considerable energy to function, and while they rely primarily on glucose, they have a relatively low rate of glycolysis compared to other cells. This limited glycolytic activity is essential for maintaining mitochondrial health and preventing redox stress. Neurons use glucose-derived metabolites from glycolysis to fuel the tricarboxylic acid (TCA) cycle for ATP production. Disruptions in glycolysis can impair cognitive function and lead to neurodegenerative diseases, highlighting the balance that neurons maintain between glycolysis and mitochondrial energy production.

Point of Interest
- Neurons rely on limited glycolysis to prevent mitochondrial dysfunction and maintain cognitive function and metabolism.

- Increasing glycolysis in neurons causes mitochondrial abnormalities, redox stress, bioenergetic deficiencies and cognitive decline in mice.

- Restoring brain NAD⁺ or reducing mitochondrial stress reverses cognitive decline caused by excessive neuronal glycolysis.

Point of Interest
- Inhibition of indoleamine 2,3-dioxygenase 1 (IDO1), an enzyme expressed in astrocytes, restores astrocytic metabolic support and rescues synaptic plasticity and memory in preclinical models of Alzheimer's disease.

- Blocking IDO1 improves glucose metabolism and lactate production in the hippocampus and improves neuronal function in AD mice.

- IDO1 inhibition, originally developed for cancer, may treat neurodegenerative diseases such as Alzheimer's and Parkinson's by restoring brain metabolism.

Point of Interest
- Neurons perform glycolysis and rely on it to supply TCA cycle metabolites for energy production.

- Deletion of GLUT3 or PKM1 in neurons results in age-related learning deficits, altered glucose metabolism, and changes in brain volume.

- Neurons require glycolysis for normal function, with compensatory mitochondrial changes occurring when glycolysis is disrupted.

Comparison of metabolic pathway in two types of cancer cells

The dependence of OXPHOS and Glycolysis in two types of cancer cells, HeLa and HepG2, were compared based on Lactate production, ATP levels, and OCR values.

Many cancer cells produce ATP through the glycolytic pathway. On the other hand, it has been recently reported that cancer cells whose glycolytic pathway is suppressed survive by shifting their energy metabolism to OXPHOS by enhancing mitochondrial function, and the dependency of metabolic pathways differs depending on cell lines.

＜Evaluation by Lactate production and ATP levels＞

We confirmed the changes in ATP and Lactate production when ATP synthesis by OXPHOS was inhibited by Oligomycin stimulation and by 2-Deoxy-D-glucose (2-DG) in the glycolytic pathway. The results showed that HeLa cells depend on Glycolysis and HepG2 cells depend on OXPHOS to synthesize ATP.

＜Evaluation by Lactate production and ATP levels＞
When OXPHOS was inhibited in HeLa cells, ATP levels remained unchanged (①), and lactate production increased (②). This suggests that even when OXPHOS is inhibited, glycolysis can be further activated. Conversely, when glycolysis is inhibited, ATP levels decrease significantly (③), indicating that energy production depends on glycolysis. On the other hand, when OXPHOS was inhibited in HepG2 cells, lactate production increased (④), indicating that the cells attempt to compensate for energy production by enhancing glycolysis, but ATP levels still decrease (⑤). This means that even with increased glycolysis, ATP production is not sufficiently compensated. Furthermore, ATP levels decrease more when glycolysis is inhibited (⑥), suggesting that energy production in HepG2 cells depends more on OXPHOS than glycolysis.

Products in Use
   - Glycolysis/OXPHOS Assay Kit

＜Evaluation by OCR value＞

Using the same number of cells, we measured the OCR value when cellular oxygen consumption was promoted by stimulating the cells with FCCP, a mitochondrial uncoupling agent. The results showed that HepG2 cells had higher OCR values than HeLa cells, suggesting a greater dependence on OXPHOS, correlating with the results obtained from ATP level and Lactate production.

Products in Use
   - Extracellular OCR Plate Assay Kit

Metabolic alterations play a critical role in the development and progression of diseases such as cancer, neurodegeneration and senescence. In cancer, altered metabolism supports rapid cell growth and survival, often through the Warburg effect, where cancer cells rely heavily on glycolysis even in the presence of oxygen. Neurodegenerative diseases such as Alzheimer's and Parkinson's are associated with dysregulated energy metabolism leading to impaired neuronal function and increased oxidative stress. Senescence, the process of cellular aging, involves metabolic reprogramming that contributes to the cessation of cell division and the secretion of pro-inflammatory factors, affecting tissue homeostasis and promoting age-related diseases.

Point of Interest
- IL-4 and tumor media upregulate phosphoglycerate dehydrogenase (PHGDH), promoting immunosuppressive M2 macrophage activation and proliferation.

- PHGDH-mediated serine biosynthesis activates mTORC1 signaling and maintains M2-like macrophages.

- Loss of PHGDH disrupts metabolism and shifts M2-like tumor-associated macrophages to an M1-like phenotype, suggesting that PHGDH inhibition may suppress tumor progression.

Point of Interest
- Metabolic and genotoxic stress promotes lysosomal cholesterol efflux and NAD⁺ depletion in macrophages via LXR/CD38 signaling.

- Cholesterol-mediated NAD+ depletion induces macrophage senescence, contributing to age-related macular degeneration (AMD) and neurodegeneration.

- NAD⁺ augmentation and senolytics reverse macrophage senescence, preventing AMD and neurodegeneration.

Point of Interest
- Defective myeloid responses in late-onset AD are associated with aging and triggering receptor expressed on myeloid cells 1 (TREM1), a pro-inflammatory factor, influences cognitive decline.

- The age-related increase in TREM1 suppresses glucose metabolism, particularly the production of ribose-5P, which is essential for both glycolysis and the biosynthesis of purines, pyrimidines and NAD⁺, leading to disruption of homeostatic microglial metabolism and immune function.

- TREM1 deficiency prevents age-related changes in myeloid metabolism, inflammation and hippocampal memory function.

Comparison of metabolic pathway in two types of cancer cells

The dependence of OXPHOS and Glycolysis in two types of cancer cells, HeLa and HepG2, were compared based on Lactate production, ATP levels, and OCR values.

Many cancer cells produce ATP through the glycolytic pathway. On the other hand, it has been recently reported that cancer cells whose glycolytic pathway is suppressed survive by shifting their energy metabolism to OXPHOS by enhancing mitochondrial function, and the dependency of metabolic pathways differs depending on cell lines.

＜Evaluation by Lactate production and ATP levels＞

We confirmed the changes in ATP and Lactate production when ATP synthesis by OXPHOS was inhibited by Oligomycin stimulation and by 2-Deoxy-D-glucose (2-DG) in the glycolytic pathway. The results showed that HeLa cells depend on Glycolysis and HepG2 cells depend on OXPHOS to synthesize ATP.

＜Evaluation by Lactate production and ATP levels＞
When OXPHOS was inhibited in HeLa cells, ATP levels remained unchanged (①), and lactate production increased (②). This suggests that even when OXPHOS is inhibited, glycolysis can be further activated. Conversely, when glycolysis is inhibited, ATP levels decrease significantly (③), indicating that energy production depends on glycolysis. On the other hand, when OXPHOS was inhibited in HepG2 cells, lactate production increased (④), indicating that the cells attempt to compensate for energy production by enhancing glycolysis, but ATP levels still decrease (⑤). This means that even with increased glycolysis, ATP production is not sufficiently compensated. Furthermore, ATP levels decrease more when glycolysis is inhibited (⑥), suggesting that energy production in HepG2 cells depends more on OXPHOS than glycolysis.

Products in Use
   - Glycolysis/OXPHOS Assay Kit

＜Evaluation by OCR value＞

Using the same number of cells, we measured the OCR value when cellular oxygen consumption was promoted by stimulating the cells with FCCP, a mitochondrial uncoupling agent. The results showed that HepG2 cells had higher OCR values than HeLa cells, suggesting a greater dependence on OXPHOS, correlating with the results obtained from ATP level and Lactate production.

Products in Use
   - Extracellular OCR Plate Assay Kit

Lipid droplets (LDs) play an important role in several diseases, including cancer, neurodegeneration and cellular senescence. In cancer, LD accumulation is often observed and can support rapid cell proliferation by providing essential lipids for membrane synthesis and energy production. Neurodegenerative diseases such as Alzheimer's and Parkinson's show aberrant lipid metabolism, with excessive LDs contributing to neuroinflammation and neuronal dysfunction. In cellular senescence, lipid droplets sequester polyunsaturated fatty acids to prevent ferroptosis, a type of cell death, thereby impacting aging and age-related diseases.

Point of Interest
- T cells in the solid tumor microenvironment (TME) must catabolize lipids via mitochondrial fatty acid oxidation (FAO) to provide energy during nutrient stress.

- The solid TME activates acetyl-coenzyme A carboxylase (ACC) in T cells, causing  lipid droplet accumulation in tumor-infiltrating T cells, which inhibits FAO.

- Restricting ACC activity rewires T cell metabolism to maintain energy during TME stress.

Point of Interest
- Cell cycle arrest suppresses ferroptosis by forming diacylglycerol acyltransferase (DGAT)–dependent lipid droplet to sequester polyunsaturated fatty acids (PUFAs).

- DGAT inhibition reshuffles PUFAs to phospholipids, which resensitizes arrested cells to ferroptosis.

- Some slow-cycling antimitotic drug–resistant cancer cells, such as 5-fluorouracil–resistant cells, have accumulation of lipid droplets and combined treatment with ferroptosis inducers and DGAT inhibitors effectively suppresses their growth.

Point of Interest
- Senescent glia in aging Drosophila brains originate from neuronal mitochondrial dysfunction and express AP1, a senescence-associated transcription factor. 

- AP1+ senescent glia cause lipid accumulation in non-senescent glia and increase senescence markers.

- Targeting AP1 in senescent glia extends lifespan, but increases oxidative damage in the brain and neuronal mitochondrial function remains poor.

Hepatotoxicity test of drug-induced lipidosis using high-content imaging

Propranolol (a sympathetic β-receptor blocker) was added to a human hepatocellular carcinoma cell line (HepG2 cells), and changes in lipid droplets were observed under a fluorescence microscope. The accumulation of lipid droplets was analyzed by measuring the number, area, and fluorescence intensity of lipid droplets from the acquired microscopic images.

＜Lipid droplet imaging data＞

  Nucleus (blue: Hoechst33342 ): Ex 385 nm, Em 460 nm  
  Lipid droplet (green: Lipi-Green): Ex 475 nm, Em 535nm

HepG2 cells were treated with propranolol 0, 10, or 30 μmol/l, lipid droplets were stained with Lipi-Green and nuclei with Hoechst 33342 and observed using a fluorescence microscope (Ti2-E inverted microscope). The results showed that lipid droplets increased in a propranolol concentration-dependent manner.

Related Products
   - Lipi-Blue/ Green/ Red/ Deep Red
   - Lipid Droplet Assay Kit - Blue / Deep Red

＜Analysis of lipid droplet accumulation relative to drug treatment concentration＞

High Content Analysis (HCA) microscope system
(Nikon Corporation https://www.microscope.healthcare.nikon.com/)

For details of staining and analysis methods, please refer to "APPLICATION NOTE: Hepatotoxicity test of drug-induced lipidosis using high-content imaging" by Nikon Corporation.

From the fluorescence images obtained, the accumulation of lipid droplets per cell was analyzed by measuring cell number from nuclei and area, number, and fluorescence intensity from lipid droplets. The results showed that the number and area of lipid droplets increased in a propranolol concentration-dependent manner, with lipid droplets forming significantly under concentration conditions of 20 μmol/l or higher. The DS-Qi2 camera, which can capture a wide range of cellular areas in a single shot, was used for imaging, and the EDF module of NIS-Elements software, which can acquire focused images of all lipid droplets, was used for analysis, enabling quantitative analysis with highly reliable statistical data. The EDF module of the NIS-Elements software allows for the acquisition of focused images of all lipid droplets.

Lactate, best known as a metabolite in the body, plays several key roles. It is primarily produced during anaerobic metabolism, such as the Warburg effect in cancer cells, as a byproduct of the conversion of glucose to energy when oxygen levels are low. Beyond its role in energy metabolism, lactate acts as a signaling molecule, influencing various cellular processes such as gene expression and inflammation regulation. In addition, recent research suggests that lactate may serve as an alternative energy source for the brain and other tissues, challenging the traditional view of lactate as a waste product.

Point of Interest
- MRE11 undergoes lactylation by CBP in response to DNA damage, a process that is dependent on ATM.

- The lactylation of MRE11 facilitates DNA end breakage and homologous recombination repair.

- Elevated lactate levels in cancer cells lead to the lactylation of MRE11, contributing to chemoresistance.

- Targeting the lactylation of MRE11 can increase the sensitivity of cancer cells to chemotherapy.

Point of Interest
- Lactate promotes the expression of hepatic hepcidin via the cAMP-PKA-Smad signaling pathway.

- Lactate directly binds to soluble adenylyl cyclase, enhancing its enzymatic activity.

- Lactate modulates iron homeostasis by inducing the expression of hepatic hepcidin.

Point of Interest
- Lactate acts as a mitochondrial messenger, promoting the shift of ATP production to the mitochondria.

- It activates the electron transport chain (ETC) independently of its own metabolism.

- Activation of the ETC by lactate results in increased pyruvate oxidation and enhanced lactate utilization.

- The mitochondrial ETC can sense the availability of lactate and recognize it as a potential nutrient.

Comparison of metabolic pathway in two types of cancer cells

The dependence of OXPHOS and Glycolysis in two types of cancer cells, HeLa and HepG2, were compared based on Lactate production, ATP levels, and OCR values.

Many cancer cells produce ATP through the glycolytic pathway. On the other hand, it has been recently reported that cancer cells whose glycolytic pathway is suppressed survive by shifting their energy metabolism to OXPHOS by enhancing mitochondrial function, and the dependency of metabolic pathways differs depending on cell lines.

＜Evaluation by Lactate production and ATP levels＞

We confirmed the changes in ATP and Lactate production when ATP synthesis by OXPHOS was inhibited by Oligomycin stimulation and by 2-Deoxy-D-glucose (2-DG) in the glycolytic pathway. The results showed that HeLa cells depend on Glycolysis and HepG2 cells depend on OXPHOS to synthesize ATP.

＜Evaluation by Lactate production and ATP levels＞
When OXPHOS was inhibited in HeLa cells, ATP levels remained unchanged (①), and lactate production increased (②). This suggests that even when OXPHOS is inhibited, glycolysis can be further activated. Conversely, when glycolysis is inhibited, ATP levels decrease significantly (③), indicating that energy production depends on glycolysis. On the other hand, when OXPHOS was inhibited in HepG2 cells, lactate production increased (④), indicating that the cells attempt to compensate for energy production by enhancing glycolysis, but ATP levels still decrease (⑤). This means that even with increased glycolysis, ATP production is not sufficiently compensated. Furthermore, ATP levels decrease more when glycolysis is inhibited (⑥), suggesting that energy production in HepG2 cells depends more on OXPHOS than glycolysis.

Products in Use
   - Glycolysis/OXPHOS Assay Kit

＜Evaluation by OCR value＞

Using the same number of cells, we measured the OCR value when cellular oxygen consumption was promoted by stimulating the cells with FCCP, a mitochondrial uncoupling agent. The results showed that HepG2 cells had higher OCR values than HeLa cells, suggesting a greater dependence on OXPHOS, correlating with the results obtained from ATP level and Lactate production.

Products in Use
   - Extracellular OCR Plate Assay Kit

Metabolic alterations play a key role in determining the fate of cancer cells. Dysregulated metabolism, often characterized by increased glucose uptake and altered nutrient utilization, provides cancer cells with the necessary energy and building blocks necessary for rapid proliferation. In addition, metabolic reprogramming can confer resistance to apoptosis and promote tumor progression. Targeting these metabolic vulnerabilities holds promise for the development of therapeutic strategies that selectively eradicate cancer cells while sparing normal tissues. Understanding the intricate interplay between metabolism and cancer cell fate is essential for advancing personalized cancer therapies.

Point of Interest
- Analysis of long-term estrogen-deprived (LTED) ER+ breast cancer cells, a model of aromatase inhibitor resistance, revealed enhanced intracellular lipid storage.

- Lipid droplets together with peroxisomes, which are enriched and active in the LTED cells, conferred metabolic adaptability to the resistant tumors.

- This reprogramming was controlled by acetyl-CoA-carboxylase-1 (ACC1), whose targeting ACC1 selectively impaired LTED survival.

- Targeting ACC1 reduced tumor growth of resistant patient-derived xenografts.

Point of Interest
- Numb, a cell fate determinant, binds to Parkin and facilitates Parkin-mediated mitophagy.

- Disruption of mitochondrial quality control triggers a metabolic-epigenetic switch.

- Histone lactylation induces transcriptional upregulation of neuroendocrine genes.

- Dysfunction in the Numb/Parkin axis propels a neuroendocrine cell fate in cancer cells.

Point of Interest
-An accumulation of diacyl-PUFA phosphatidylcholines (PC-PUFA2s) after fatty acid or phospholipid treatments correlates with cancer cell sensitivity to ferroptosis.

-PC-PUFA2s interacted with the mitochondrial electron transport chain, generating ROS to initiate lipid peroxidation.

-Mitochondria-targeted antioxidants protected cells from PC-PUFA2-induced mtROS, lipid peroxidation, and cell death.

Comparison of metabolic pathway in two types of cancer cells

The dependence of OXPHOS and Glycolysis in two types of cancer cells, HeLa and HepG2, were compared based on Lactate production, ATP levels, and OCR values.

Many cancer cells produce ATP through the glycolytic pathway. On the other hand, it has been recently reported that cancer cells whose glycolytic pathway is suppressed survive by shifting their energy metabolism to OXPHOS by enhancing mitochondrial function, and the dependency of metabolic pathways differs depending on cell lines.

＜Evaluation by Lactate production and ATP levels＞

We confirmed the changes in ATP and Lactate production when ATP synthesis by OXPHOS was inhibited by Oligomycin stimulation and by 2-Deoxy-D-glucose (2-DG) in the glycolytic pathway. The results showed that HeLa cells depend on Glycolysis and HepG2 cells depend on OXPHOS to synthesize ATP.

＜Evaluation by Lactate production and ATP levels＞
When OXPHOS was inhibited in HeLa cells, ATP levels remained unchanged (①), and lactate production increased (②). This suggests that even when OXPHOS is inhibited, glycolysis can be further activated. Conversely, when glycolysis is inhibited, ATP levels decrease significantly (③), indicating that energy production depends on glycolysis. On the other hand, when OXPHOS was inhibited in HepG2 cells, lactate production increased (④), indicating that the cells attempt to compensate for energy production by enhancing glycolysis, but ATP levels still decrease (⑤). This means that even with increased glycolysis, ATP production is not sufficiently compensated. Furthermore, ATP levels decrease more when glycolysis is inhibited (⑥), suggesting that energy production in HepG2 cells depends more on OXPHOS than glycolysis.

Products in Use
   - Glycolysis/OXPHOS Assay Kit

＜Evaluation by OCR value＞

Using the same number of cells, we measured the OCR value when cellular oxygen consumption was promoted by stimulating the cells with FCCP, a mitochondrial uncoupling agent. The results showed that HepG2 cells had higher OCR values than HeLa cells, suggesting a greater dependence on OXPHOS, correlating with the results obtained from ATP level and Lactate production.

Products in Use
   - Extracellular OCR Plate Assay Kit

Scientists have discovered that in both basal and activated states, neuronal somata have higher levels of aerobic glycolysis and lower levels of OXPHOS than terminals. Their findings update the conventional view that neurons use OXPHOS uniformly under basal conditions and highlight the important role of somatic aerobic glycolysis in maintaining antioxidant capacity.

Aerobic glycolysis is the predominant means of glucose metabolism in neuronal somata, which protects against oxidative damage
Click here for the original article: Yao Wei, et. al., Nature Neuroscience, 2023.

Point of Interest
- Neuronal somata engage more in aerobic glycolysis and less in OXPHOS compared to axon terminals, both at rest and when activated.
- The enzyme pyruvate kinase 2 (PKM2) is primarily found in neuronal somata, not terminals.
- Deleting Pkm2 in mice shifts somatic metabolism from aerobic glycolysis to OXPHOS.
- This shift causes oxidative damage and loss of dopaminergic neurons.

Comparison of metabolic pathway in two types of cancer cells

The dependence of OXPHOS and Glycolysis in two types of cancer cells, HeLa and HepG2, were compared based on Lactate production, ATP levels, and OCR values.

Many cancer cells produce ATP through the glycolytic pathway. On the other hand, it has been recently reported that cancer cells whose glycolytic pathway is suppressed survive by shifting their energy metabolism to OXPHOS by enhancing mitochondrial function, and the dependency of metabolic pathways differs depending on cell lines.

＜Evaluation by Lactate production and ATP levels＞

We confirmed the changes in ATP and Lactate production when ATP synthesis by OXPHOS was inhibited by Oligomycin stimulation and by 2-Deoxy-D-glucose (2-DG) in the glycolytic pathway. The results showed that HeLa cells depend on Glycolysis and HepG2 cells depend on OXPHOS to synthesize ATP.

＜Evaluation by Lactate production and ATP levels＞
When OXPHOS was inhibited in HeLa cells, ATP levels remained unchanged (①), and lactate production increased (②). This suggests that even when OXPHOS is inhibited, glycolysis can be further activated. Conversely, when glycolysis is inhibited, ATP levels decrease significantly (③), indicating that energy production depends on glycolysis. On the other hand, when OXPHOS was inhibited in HepG2 cells, lactate production increased (④), indicating that the cells attempt to compensate for energy production by enhancing glycolysis, but ATP levels still decrease (⑤). This means that even with increased glycolysis, ATP production is not sufficiently compensated. Furthermore, ATP levels decrease more when glycolysis is inhibited (⑥), suggesting that energy production in HepG2 cells depends more on OXPHOS than glycolysis.

Products in Use
   - Glycolysis/OXPHOS Assay Kit

＜Evaluation by OCR value＞

Using the same number of cells, we measured the OCR value when cellular oxygen consumption was promoted by stimulating the cells with FCCP, a mitochondrial uncoupling agent. The results showed that HepG2 cells had higher OCR values than HeLa cells, suggesting a greater dependence on OXPHOS, correlating with the results obtained from ATP level and Lactate production.

Products in Use
   - Extracellular OCR Plate Assay Kit

Scientists have found that deficiency of two key glycolytic enzymes, Glut1 (glucose transporter 1) and Gpi1 (glucose-6-phosphate isomerase 1), enhanced tumor cell killing by cytotoxic T cells. Genetic and pharmacologic inactivation of Glut1 sensitizes tumors to anti-tumor immunity and synergizes with anti-PD-1 therapy through the TNF-α pathway.

Tumor aerobic glycolysis confers immune evasion through modulating sensitivity to T cell-mediated bystander killing via TNF-α
Click here for the original article: Lijian Wu, et. al., Cell Metabolism, 2023.

Point of Interest
- The intrinsic glycolysis pathway within tumors confers resistance to killing by T cells.
- Depletion of Glut1 sensitizes tumor cells to cell death induced by TNF-α through the augmentation of reactive oxygen species (ROS).
- Glut1 and Glut3 exhibit differential expression in tumor cells compared to immune cells.
- Inhibiting Glut1 enhances the anti-tumor immune response in murine models.

Metabolic shift to glycolysis in senescenct cells

＜Evaluation by Lactate production and ATP levels＞

NAD(+) levels decline during the aging process, causing defects in nuclear and mitochondrial functions and resulting in many age-associated pathologies*. Here, we try to redemonstrate this phenomenon in the doxorubicin (DOX)-induced cellular senescence model with a comprehensive analysis of our products.

*S. Imai, et al., Trends Cell Biol, 2014, 24, 464-471

Products in Use
① DNA Damage Detection Kit - γH2AX
② Cellular Senescence Detection Kit - SPiDER-βGal
③ NAD/NADH Assay Kit-WST
④ JC-1 MitoMP Detection Kit
⑤ Glycolysis/OXPHOS Assay Kit、Lactate Assay Kit-WST

Scientists have found that lysosomal cystine deprivation, unlike general amino acid starvation, triggers ATF4 induction at the transcriptional level, but cytosolic cysteine deprivation does not. Blocking lysosomal cystine efflux diminishes ATF4 induction and enhances sensitivity to ferroptosis. Therefore, intracellular nutrient reprogramming could potentially induce selective ferroptosis in cancer cells without causing systemic starvation.

Lysosomal cystine governs ferroptosis sensitivity in cancer via cysteine stress response
Click here for the original article: Robert V. Swanda, et. al., Molecular Cell, 2023.

Point of Interest
- Depletion of cysteine induces adaptive ATF4 expression at the transcriptional level.
- A shortage of cystine in lysosomes stimulates ATF4 expression through the AhR signaling pathway.
- A weakened cysteine stress response increases sensitivity to ferroptosis during cysteine deprivation.
- CysRx promotes cancer cell ferroptosis through intracellular nutrient reprogramming..

Lysosomal Acidic pH Detection Kit-Green/Red 
Lysosomal Acidic pH Detection Kit-Green/Deep Red 

Lysosomal Function and Mitochondrial ROS

CCCP and Antimycin are recognized inducers of mitochondrial ROS, linked to the loss of mitochondrial membrane potential. Recent studies have shown that CCCP induces not only mitochondrial ROS but also lysosomal dysfunction. To observe mitochondrial ROS, HeLa cells were labeled with MitoBright ROS Deep Red for Mitochondrial Superoxide Detection, and the lysosomal mass and pH were independently detected with LysoPrime Green and pHLys Red. Co-staining with MitoBright ROS and Lysosomal dyes revealed that CCCP, unlike Antimycin, triggers concurrent lysosomal neutralization and mitochondrial ROS induction.

Reference: Benjamin S Padman, et. al., Autophagy (2013)

Products in Use
   - LysoPrime Green
   - pHLys Red
   - Lysosomal Acidic pH Detection Kit
   - MitoBright ROS Deep Red - Mitochondrial Superoxide Detection

Lysosomal Function and Iron Homeostasis

Recent reports suggest that lysosomal neutralization can result in iron depletion, consequently leading to the disruption of cell viability. To verify this, HeLa cells were labeled with FerroOrange for Fe²⁺ detection, and the lysosomal mass and pH were separately detected with LysoPrime DeepRed and pHLys Green (a product currently under development). Co-staining with FerroOrange and Lysosomal dyes demonstrated that Bafilomycin A1 (Baf. A1), an inhibitor of lysosomal acidification, causes iron depletion consistent with the findings reported in the article. Interestingly, the iron chelator, Deferiprone (DFP), did not impact lysosomal pH, suggesting that lysosomal function plays a key role in managing iron homeostasis.

Reference: Ross A Weber, et. al., Mol Cell (2020)

Products in Use
   - FerroOrange
   - pHLys Green*
   - LysoPrime Deep Red

*pHLys Green is a component of "Lysosomal Acidic pH Detection Kit-Green/Deep Red".

Scientists have found that neuroinflammatory lesions in the mouse spinal cord cause widespread and persistent axonal ATP deficiency that precedes mitochondrial oxidation and calcium overload. Notably, viral overexpression of individual TCA enzymes can ameliorate axonal energy deficits in neuroinflammatory lesions. This suggests that TCA cycle dysfunction in the common neuroinflammatory disease multiple sclerosis may be amenable to therapy.

Targeting the TCA cycle can ameliorate widespread axonal energy deficiency in neuroinflammatory lesions
Click here for the original article: Yi-Heng Tai, et. al., Nature Metabolism, 2023.

Point of Interest
- Neuroinflammatory lesions in mice cause long-lasting axonal ATP deficiency.
- This deficiency precedes mitochondrial oxidation and calcium overload.
- It's linked to tricarboxylic acid (TCA) cycle enzyme imbalances.
- Correcting TCA enzyme levels can potentially treat these energy deficits.

Comparison of metabolic pathway in two types of cancer cells

The dependence of OXPHOS and Glycolysis in two types of cancer cells, HeLa and HepG2, were compared based on Lactate production, ATP levels, and OCR values.

Many cancer cells produce ATP through the glycolytic pathway. On the other hand, it has been recently reported that cancer cells whose glycolytic pathway is suppressed survive by shifting their energy metabolism to OXPHOS by enhancing mitochondrial function, and the dependency of metabolic pathways differs depending on cell lines.

＜Evaluation by Lactate production and ATP levels＞

We confirmed the changes in ATP and Lactate production when ATP synthesis by OXPHOS was inhibited by Oligomycin stimulation and by 2-Deoxy-D-glucose (2-DG) in the glycolytic pathway. The results showed that HeLa cells depend on Glycolysis and HepG2 cells depend on OXPHOS to synthesize ATP.

＜Evaluation by Lactate production and ATP levels＞
When OXPHOS was inhibited in HeLa cells, ATP levels remained unchanged (①), and lactate production increased (②). This suggests that even when OXPHOS is inhibited, glycolysis can be further activated. Conversely, when glycolysis is inhibited, ATP levels decrease significantly (③), indicating that energy production depends on glycolysis. On the other hand, when OXPHOS was inhibited in HepG2 cells, lactate production increased (④), indicating that the cells attempt to compensate for energy production by enhancing glycolysis, but ATP levels still decrease (⑤). This means that even with increased glycolysis, ATP production is not sufficiently compensated. Furthermore, ATP levels decrease more when glycolysis is inhibited (⑥), suggesting that energy production in HepG2 cells depends more on OXPHOS than glycolysis.

Products in Use
   - Glycolysis/OXPHOS Assay Kit

＜Evaluation by OCR value＞

Using the same number of cells, we measured the OCR value when cellular oxygen consumption was promoted by stimulating the cells with FCCP, a mitochondrial uncoupling agent. The results showed that HepG2 cells had higher OCR values than HeLa cells, suggesting a greater dependence on OXPHOS, correlating with the results obtained from ATP level and Lactate production.

Products in Use
   - Extracellular OCR Plate Assay Kit

Scientists have discovered that disablement of fatty acid oxidation in cardiomyocytes improves resistance to hypoxia and stimulates cardiomyocyte proliferation, allowing heart regeneration after ischaemia–reperfusion injury. This process involves epigenetic inhibition of cardiomyocyte maturation via activation of the alpha-ketoglutarate-dependent lysine demethylase KDM5.

Inhibition of fatty acid oxidation enables heart regeneration in adult mice
Click here for the original article: Xiang Li, et. al., Nature, 2023.

Point of Interest
- Cardiomyocytes undergo postnatal maturation which limits heart regeneration.
- Disabling fatty acid oxidation in these cells promotes their proliferation and aids heart repair after injury.  
- This process is linked to changes in energy metabolism and the activation of KDM5, altering gene expression.  
- Reversing metabolic maturation can rejuvenate cardiomyocytes and facilitate heart healing.

Fatty acid starvation induced by uptake inhibitor evoke reprogramming of cellular metabolism 

Mitochondrial fatty acid β-oxidation and oxidative phosphorylation (OXPHOS) are crucial biochemical processes that metabolize fats and sugars to produce ATP, the cell's primary energy source. In this section, we underscored the significance of fatty acid starvation and energy pathways, with an emphasis on the fatty acid uptake inhibitor, FATP2. Here are the key findings from our experiments conducted on HeLa cells:

・Inhibition of fatty acid uptake results in reduced cell proliferation, though it does not lead to cell death. This was determined through the use of a Cell Counting Kit-8 and Fatty Acid Uptake Kit (Image 1).

・Fatty acid starvation shifts cellular metabolism from OXPHOS to glycolysis, as indicated by the Glycolysis/JC-1 MitoMP Assay Kit. (Image 2)

・When fatty acid uptake is inhibited, a compensatory increase in glucose and glutamine uptake occurs to preserve cell viability, as observed using the Glucose Assay Kit and Glutamine Assay Kit. (Image 3)

Products in Use
for Fatty Acid Uptake Assay
  ① Fatty Acid Uptake Assay Kit

for Cell Proliferation/Cytotoxicity Assay
  ② Cell Counting Kit-8

for Glycolysis Assay and Mitochondrial Membrane Potential Detection
  ③ Glycolysis/JC-1 MitoMP Assay Kit

for Glucose and Glutamine Consumption Assay
  ④ Glucose Assay Kit

  ⑤ Glutamine Assay Kit

In recent years, the discovery of several novel lipid-mitochondria axis in metabolism has attracted much attention. A link between acylcarnitine accumulation and lipid intolerance in skeletal muscle mitochondria is tightly linked to poor health. In other breakthroughs, decreased expression in fatty acid transporters and β-oxidation enzymes causes fatty acid buildup in lipid droplets. Research also uncovers that knocking out a vital enzyme in mitochondrial fatty acid oxidation astrocytes leads to cognitive impairment.

Pyruvate-supported flux through medium-chain ketothiolase promotes mitochondrial lipid tolerance in cardiac and skeletal muscles
Click here for the original article: Timothy R. Koves, et. al., Cell Metabolism, 2023.

Arf1 coordinates fatty acid metabolism and mitochondrial homeostasis
Click here for the original article: Ludovic Enkler, et. al., Nature Cell Biology, 2023.

Fatty acid oxidation organizes mitochondrial supercomplexes to sustain astrocytic ROS and cognition
Click here for the original article: Brenda Morant-Ferrando , et. al., Nature Metabolism, 2023.

Point of Interest
- The metabolic pathway of long-chain fatty acid oxidation is susceptible to bottlenecks.
- Bottlenecks in the oxidation of fat can lead to CoA trapping and a decrease in respiratory efficiency.
- The reverse flux of pyruvate-derived acetyl CoA through the medium-chain ketothiolase pathway can regenerate free CoA.
- High levels of medium-chain ketothiolase in the heart and red muscles enhance mitochondrial tolerance to lipids.

Point of Interest
- Expression of fatty acid transporters and the key enzyme in β-oxidation decreased in cells with a hyperactive Arf1 mutant, causing fatty acids to accumulate in lipid droplets.
- As a result, mitochondria fragmented and ATP synthesis declined.
- Genetic and drug-induced reduction of fatty acids mimicked the mitochondrial effects of the arf1 mutant.
- Although β-oxidation occurs in both mitochondria and peroxisomes in mammals, Arf1's role in fatty acid metabolism remains conserved.

Point of Interest
- Knockout of CPT1A, a key enzyme of mitochondrial fatty acid oxidation, leads to cognitive impairment in adult mouse astrocytes.
- Decreased fatty acid oxidation alters astrocytic pyruvate metabolism and reduces reactive oxygen species formation.
- Astrocytes naturally metabolize fatty acids to maintain the mitochondrial respiratory chain in an energetically inefficient, disassembled conformation.
- This disassembled conformation helps in preserving the signaling of reactive oxygen species and supports cognitive performance.

Fatty acid starvation induced by uptake inhibitor evoke reprogramming of cellular metabolism 

Mitochondrial fatty acid β-oxidation and oxidative phosphorylation (OXPHOS) are crucial biochemical processes that metabolize fats and sugars to produce ATP, the cell's primary energy source. In this section, we underscored the significance of fatty acid starvation and energy pathways, with an emphasis on the fatty acid uptake inhibitor, FATP2. Here are the key findings from our experiments conducted on HeLa cells:

・Inhibition of fatty acid uptake results in reduced cell proliferation, though it does not lead to cell death. This was determined through the use of a Cell Counting Kit-8 and Fatty Acid Uptake Kit (Image 1).

・Fatty acid starvation shifts cellular metabolism from OXPHOS to glycolysis, as indicated by the Glycolysis/JC-1 MitoMP Assay Kit. (Image 2)

・When fatty acid uptake is inhibited, a compensatory increase in glucose and glutamine uptake occurs to preserve cell viability, as observed using the Glucose Assay Kit and Glutamine Assay Kit. (Image 3)

Products in Use
for Fatty Acid Uptake Assay
  ① Fatty Acid Uptake Assay Kit

for Cell Proliferation/Cytotoxicity Assay
  ② Cell Counting Kit-8

for Glycolysis Assay and Mitochondrial Membrane Potential Detection
  ③ Glycolysis/JC-1 MitoMP Assay Kit NEW

for Glucose and Glutamine Consumption Assay
  ④ Glucose Assay Kit

  ⑤ Glutamine Assay Kit

Loss of UCHL1 rescues the defects related to Parkinson’s disease by suppressing glycolysis
Su Jin Ham, et. al., Sci Adv, 7 (2021)

   Point of Interest
   - The loss of UCHL1 regulates glucose metabolism and rescues the defects related to PD. 
   - Specific glycolytic metabolites are decreased and an energy-dependent mitophagy pathway is induced by inhibition of UCHL1.
   - UCHL1 plays a critical role in regulating mitophagy independent of the PINK1-Parkin pathway and PKM is the responsible target of UCHL1 in controlling mitophagy and PD pathogenesis.

Fatty acid starvation induced by uptake inhibitor evoke reprogramming of cellular metabolism 

Mitochondrial fatty acid β-oxidation and oxidative phosphorylation (OXPHOS) are crucial biochemical processes that metabolize fats and sugars to produce ATP, the cell's primary energy source. In this section, we underscored the significance of fatty acid starvation and energy pathways, with an emphasis on the fatty acid uptake inhibitor, FATP2. Here are the key findings from our experiments conducted on HeLa cells:

・Inhibition of fatty acid uptake results in reduced cell proliferation, though it does not lead to cell death. This was determined through the use of a Cell Counting Kit-8 and Fatty Acid Uptake Kit (Image 1).

・Fatty acid starvation shifts cellular metabolism from OXPHOS to glycolysis, as indicated by the Glycolysis/JC-1 MitoMP Assay Kit. (Image 2)

・When fatty acid uptake is inhibited, a compensatory increase in glucose and glutamine uptake occurs to preserve cell viability, as observed using the Glucose Assay Kit and Glutamine Assay Kit. (Image 3)

Products in Use
for Fatty Acid Uptake Assay
  ① Fatty Acid Uptake Assay Kit

for Cell Proliferation/Cytotoxicity Assay
  ② Cell Counting Kit-8

for Glycolysis Assay and Mitochondrial Membrane Potential Detection
  ③ Glycolysis/JC-1 MitoMP Assay Kit NEW

for Glucose and Glutamine Consumption Assay
  ④ Glucose Assay Kit

  ⑤ Glutamine Assay Kit

Large neutral amino acid levels tune perinatal neuronal excitability and survival
Lisa Knaus, et. al., Cell, 186, 1-18 (2023)

   Point of Interest
   - Metabolic reprogramming of the cerebral cortex throughout development
   - Essential role of large neutral amino acids in perinatal neuronal metabolic state
   - Link between Slc7a5 deletion in neurons, branched-chain amino acids, and lipid metabolism
   - Impact of reduced large neutral amino acid levels on neuronal excitability and survival

The metabolic shift to glycolysis under the treatment of fatty acid transporter inhibitors

Products in Use
for Fatty Acid Uptake Assay
  ① Fatty Acid Uptake Assay Kit

for Cell Proliferation/Cytotoxicity Assay
  ② Cell Counting Kit-8

for Glycolysis Assay
  ③ Glucose Assay Kit-WST
  ④ Lactate Assay Kit-WST

Related Products
for Glycolysis and OXPHOS Assay
  - Glycolysis/OXPHOS Assay Kit

for OXPHOS Assay
  - Extracellular OCR Plate Assay Kit

Lactylome analysis suggests lactylation-dependent mechanisms of metabolic adaptation in hepatocellular carcinoma
Zijian Yang, et. al., Nature Metabolism, 5, 61-79 (2023)

   Point of Interest
   - Lactate drives the posttranslational modification, lysine lactylation (Kla), on core histones but other targets remain unclear.
   - Lactylome and proteome analysis of the tumors and adjacent livers reveals 99.8% of Kla sites are on non-histone proteins. 
   - Kla affects enzymes involved in metabolic pathways, including nucleotide metabolism.
   - Lactylation at K28 inhibits adenylate kinase 2, facilitating the proliferation and metastasis of HCC cells.

Metabolic shift to glycolysis in senescenct cells

	NAD(+) levels decline during the aging process, causing defects in nuclear and mitochondrial functions and resulting in many age-associated pathologies*. Here, we try to redemonstrate this phenomenon in the doxorubicin (DOX)-induced cellular senescence model with a comprehensive analysis of our products. *S. Imai, et al., Trends Cell Biol, 2014, 24, 464-471

Products in Use
① DNA Damage Detection Kit - γH2AX

② Cellular Senescence Detection Kit - SPiDER-βGal

③ NAD/NADH Assay Kit-WST

④ JC-1 MitoMP Detection Kit

⑤ Glycolysis/OXPHOS Assay Kit、Lactate Assay Kit-WST

• - Using ¹³C-tracer analysis, the activity of glutamine-to-fumarate, citrate-to-succinate, and glutamine-to-aspartate routes was identified in the nuclei.
• - Proximity labeling mass spectrometry revealed a spatial vicinity of the involved enzymes with core nuclear proteins.
• - 2-oxoglutarate dehydrogenase, which produces succinyl-CoA, also localizes at the nucleus in mouse embryonic stem cells.
• - The nuclear localization of this enzyme changed from pluripotency to a differentiated state with accompanying changes in the nuclear protein succinylation.

• - iNs from patients with AD express cancer-associated PKM2
• - PKM2 facilitates Warburg-effect-like glycolytic reprogramming of old neurons
• - Nuclear PKM2 associates with STAT3 and HIF1α to promote neuronal fate loss in AD iNs
• - Modulation of PKM2 with shikonin restores healthy neuronal features

• - Adipocytes transfer mitochondria to tissue-specific networks of cells in fat
• - Diet-induced obesity, but not aging, impairs mitochondria transfer to macrophages
• - Dietary long-chain fatty acids inhibit mitochondria uptake by macrophages
• - Macrophages limit the release of adipocyte mitochondria into blood