Science Note: Senescence

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.

Analysis of Lysosomal Mass and pH change 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
   > More about Lysosomal Function Analysis