Ferroptosis is a form of programmed cell death characterized by the accumulation of lipid peroxides to lethal levels and is distinct from other forms of cell death such as apoptosis, necroptosis, and autophagy. In the context of cancer, ferroptosis may act as a tumor suppressor mechanism, as cancer cells often have an increased susceptibility to ferroptosis due to their altered metabolism and increased levels of reactive oxygen species (ROS). Therapeutically, inducing ferroptosis in cancer cells has emerged as a promising strategy for cancer treatment, particularly for tumors that are resistant to traditional therapies such as chemotherapy and radiation. In addition, understanding the specific vulnerabilities of cancer cells to ferroptosis may aid in the design of targeted therapies that exploit these weaknesses, providing a potential avenue to overcome drug resistance and improve patient outcomes.

Dietary restriction of cysteine and methionine sensitizes gliomas to ferroptosis and induces alterations in energetic metabolism
Click here for the original article: Pavan S. Upadhyayula, et. al., Nature Communications, 2023.

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.

Mitochondria regulate intracellular coenzyme Q transport and ferroptotic resistance via STARD7
Click here for the original article: Soni Deshwal et. al., Nature Cell Biology, 2023.

Point of Interest
- Cysteine and methionine deprivation (CMD) can synergize with the GPX4 inhibitor RSL3 to increase ferroptotic cell death and lipid peroxidation.

- A cysteine-depleted, methionine-restricted diet can improve the therapeutic response to RSL3 and prolong survival in a syngeneic orthotopic murine glioma model.

- This CMD diet profoundly alters in vivo metabolome, proteome, and lipidome.

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.

Point of Interest
- The rhomboid protease PARL cleaves STARD7, allowing its dual localization to the mitochondrial intermembrane space and cytosol.

- The mitochondrial STARD7 supports coenzyme Q synthesis, promotes oxidative phosphorylation, and maintains cristae morphology.

- Its cytosolic counterpart facilitates the transport of coenzyme Q to the plasma membrane and protects against ferroptosis.

- Overexpression of cytosolic STARD7 increases the resistance of cells to ferroptosis and reduces the availability of coenzyme Q in the mitochondria.

Autophagy is a cellular process that degrades and recycles damaged organelles, misfolded proteins, and other cellular debris, and is critical for maintaining cellular homeostasis. Dysregulated autophagy is often observed in the context of neurodegeneration, where either excessive or insufficient autophagic activity can contribute to the accumulation of toxic proteins and organelle dysfunction that are hallmarks of neurodegenerative diseases. Enhancing autophagy has been shown to alleviate symptoms and pathology in models of diseases such as Alzheimer's, Parkinson's, and Huntington's, suggesting a protective role against neurodegeneration. Conversely, excessive autophagy can lead to cell death, suggesting that a delicate balance is required for neuronal health and highlighting the complexity of targeting autophagy as a therapeutic strategy for neurodegenerative diseases.

Point of Interest
- Inhibiting microglial autophagy leads to the disengagement of microglia from amyloid plaques, exacerbating neuropathology in mice with Alzheimer's Disease (AD).

- An autophagy deficiency fosters the development of senescent microglia.

- Pharmacological removal of autophagy-deficient senescent microglia can alleviate neuropathology in mice with AD.

Point of Interest
- Activated microglia mediate non-cell-autonomous inhibition of neuronal autophagy.

- CCL-5, CCL-4, and CCL-3 derived from microglia activate the neuronal CCR5 receptor, leading to the inhibition of neuronal autophagy.

- Levels of CCR5, CCL-3, CCL-4, and CCL-5 are elevated in the brains of mice with Huntington’s disease and tauopathy.

- Inhibiting CCR5, either pharmacologically or genetically, ameliorates neurodegeneration in mice.

Point of Interest
- The translocator protein (TSPO) is pivotal for respiratory metabolism and energy supply in microglia.

- Hexokinase-2 (HK) affects glycolytic metabolism and phagocytosis through its interaction with mitochondria.

- Mitochondrial HK influences glycolysis and inflammation, and its displacement improves phagocytosis in TSPO-deficient microglia.

- Alzheimer’s beta-amyloid drastically stimulated mitochondrial HK recruitment in cultured microglia, which may contribute to microglial dysfunction in Alzheimer’s disease.

Autolysosome
(DALGreen)

＜Cellular Landscape: autophagy-related pathway＞

Link to each product (red at left figure)

Autophagy Detection
 Autophagic Flux Assay Kit - First-choice product
 DAPRed - Autophagosome detection
 DAPGreen - Autophagosome detection
 DALGreen - Autolysosome detection

Mitophagy Detection
 Mitophagy Detection Kit and Mtphagy Dye

Mitochondrial Membrane Potential Detection
 JC-1 / MT-1

Lysosomal Analysis
 Lysosomal Acidic pH Detection Kit
                   Green/Red, Green/Deep Red

Endocytosis Detection
 ECGreen-Endocytosis Detection
 AcidSensor Labeling Kit

Exosome
 Exosome Isolation Kit
 Exosome Membrane Labeling Kit
                          Green / Red / Deep Red

Senescence is a cellular process that results in the cessation of cell division, often serving as a protective mechanism against the proliferation of damaged cells, including potential cancer cells. This process is intricately regulated by numerous factors including, but not limited to, tumor suppressor genes, DNA damage response (DDR) pathways, and various signaling molecules. In addition, the senescence-associated secretory phenotype (SASP), consisting of cytokines, growth factors, and proteases, is regulated by NF-κB and other transcription factors that influence the tissue microenvironment and impact aging and disease processes.

HKDC1, a target of TFEB, is essential to maintain both mitochondrial and lysosomal homeostasis, preventing cellular senescence
Click here for the original article: Mengying Cui, et. al., PNAS, 2023.

Point of Interest
- HKDC1, a protein involved in glycolysis, is a direct target of the transcription factor TFEB and is essential for maintaining both mitochondrial and lysosomal function.

- This activity helps avert cellular senescence, playing a vital role in maintaining cellular homeostasis.

- Beyond its role in glycolysis, HKDC1 contributes to mitophagy and lysosomal repair processes independently.

- The absence of HKDC1 may result in cellular senescence and the buildup of damaged organelles.

Point of Interest
- Vascular and hemolytic injury trigger iron accumulation, which causes senescence and promotes fibrosis.

- Senescent cells persistently accumulate iron, even after the increase in extracellular iron has subsided.

- Cells exposed to various types of senescence-inducing insults accumulate abundant ferritin-bound iron, mostly within lysosomes.
- The high levels of labile iron fuel the generation of reactive oxygen species and the SASP.  

- STK38 and GABARAPs are key regulators of this process.

- STK38 is required for lysosomal recruitment of VPS4 and GABARAPs are involved in ESCRT assembly.

- Depletion of these regulators leads to accelerated cellular senescence and shortened lifespan.

Analysis of Lysosomal Mass and pH Exchange in Senescence-induced Cells

Purpose: To investigate changes in lysosomal mass and pH in A549 cells induced to senescence by treatment with Doxorubicin (DOX).

Methods: Senescence-associated acidic β-galactosidase (SA-βGal) activity was detected using Cellular Senescence Detection Kit - SPiDER-βGal. Lysosomal mass was detected using LysoPrime Deep Red, and pH was detected using pHLys Red. Fluorescence imaging was used to observe changes in lysosomal mass and pH in senescent cells compared to non-senescent cells. The normalized fluorescence intensity of lysosomal mass and pH was also measured by a plate reader.

Results: Our findings indicate that senescence induced by DOX resulted in an increase in lysosomal mass and acidification of pH compared to non-senescent cells. The obtained results are consistent with previous reports* that demonstrated enhanced lysosomal activity in senescent cells induced by the CDK4/6 inhibitor, palbociclib. The fluorescence imaging and plate reader data both support these findings.

* Miguel Rovira, et. al., Aging Cell (2022)

＜Experimental Conditions for Microscopy＞
SA-βGal(Green):Ex = 488 nm, Em = 490 – 550 nm
Lysosomal pH (Red):Ex = 561 nm, Em = 560 – 620 nm
Lysosomal mass (Deep Red):Ex = 633 nm, Em = 640 – 700 nm

＜Experimental Conditions for Plate Reader＞
SA-βGal: Ex = 525 – 535 nm, Em = 550 – 570 nm
Lysosomal pH: Ex = 555 – 565 nm, Em = 590 – 610 nm
Lysosomal mass: Ex = 645 – 655 nm, Em = 690 – 710 nm

<Products in Use>
- Cellular Senescence Detection Kit
- Lysosomal pH and mass detection Kit
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