Journal: Biomolecules

Article Title: EET-Based Therapeutics Mitigate Sorafenib-Associated Glomerular Cell Damage

doi: 10.3390/biom15091324

Figure Lengend Snippet: 8,9-EET Analog Mitigates Sorafenib-Induced HRMCs Death by Suppressing Caspase 3/7 Activity. Caspase 3/7 activity, a marker of sorafenib-induced apoptosis, is visualized as green, fluorescent spots. An increase in green fluorescence indicates elevated caspase 3/7 activity and greater cell death, while a decrease reflects reduced apoptotic activity. HRMCs were seeded into 96-well plates for compound screening. Each plate was used to test two to three compounds, alongside matched vehicle and sorafenib (10 µM) controls. Caspase 3/7 activation was assessed using a luminescent assay, with control datasets applied uniformly across all compounds tested on the same plate. MDB-32, MDB-52a, and MDB-52b were each tested on separate 96-well plates, with individual sets of vehicle and sorafenib controls specific to each compound. In contrast, MDB-77 and MDB-78 were tested concurrently on a single 96-well plate, sharing a common set of vehicle and sorafenib controls. This design enabled consistent intraplate comparisons while maintaining compound-specific control conditions. ( a ) Treatment with the 8,9-EET analog MDB-32 led to a 20–40% reduction in sorafenib-induced caspase 3/7 activity, suggesting a protective effect against apoptosis. ( b ) Cells treated with MDB-52a in combination with 10 µM sorafenib showed significantly lower caspase 3/7 activity compared to sorafenib alone. MDB-52a reduced activity in a dose-dependent manner by 60–90%, demonstrating strong anti-apoptotic efficacy in HRMC cells. ( c ) A similar reduction in caspase 3/7 activity was observed with MDB-52b, further supporting its protective role. ( d , e ) HRMC cells treated with MDB-77 and MDB-78 also exhibited decreased caspase 3/7 activity relative to sorafenib-treated controls, indicating potential anti-apoptotic effects of these compounds.

Article Snippet: The caspase 3/7 dye was obtained from Sartorius (Bohemia, NY, USA).

Techniques: Activity Assay, Marker, Fluorescence, Activation Assay, Luminescence Assay, Control

Journal: Biomolecules

Article Title: EET-Based Therapeutics Mitigate Sorafenib-Associated Glomerular Cell Damage

doi: 10.3390/biom15091324

Figure Lengend Snippet: 8,9 EET Analog Mitigates Sorafenib-Induced Cell Death of Podocyte by Suppressing Caspase 3/7 Activity. The protective potential of 8,9-EET analogs against sorafenib-induced apoptosis in podo-cytes was evaluated by measuring caspase 3/7 activity, a key marker of programmed cell death. Apoptotic activity was visualized as green, fluorescent spots, an increased number of spots indi-cates elevated caspase 3/7 activity, while fewer spots suggest reduced apoptosis. Human podo-cytes were seeded into 96-well plates to evaluate caspase 3/7 activation in response to various test compounds. Each plate was configured to include two to three compounds, along with matched vehicle and sorafenib (10 µM) controls. A single set of control data (vehicle and sorafenib) was used for all compounds tested on the same plate to ensure consistent intra-plate comparisons. MDB-52a, MDB-52b, and RM-84 were tested concurrently on one 96-well plate, sharing a common set of vehicle and sorafenib controls. Similarly, MDB-77 and MDB-78 were tested together on a separate plate, also using a single set of vehicle and sorafenib controls for both compounds. Treatment with 8,9-EET analogs significantly reduced caspase 3/7 activity induced by sorafenib, indicating their protective effects: ( a ) MDB-52a: Co-treatment with MDB-52a and 10 µM sorafenib resulted in a marked reduction in caspase 3/7 activity compared to sorafenib alone. The effect was dose-dependent, with a reduction of approximately 50–70%. ( b ) MDB-52b: Similar protective effects were observed, with caspase 3/7 activity reduced by 40–60% in a dose-dependent manner. ( c , d ) MDB-77 and MDB-78: Both compounds decreased caspase 3/7 activity when combined with sorafenib. MDB-77 was effective at 1 and 3 µM, while MDB-78 showed minimal effect at lower doses but was effective at 10 µM. ( e ) RM-84: This analog also demonstrated a dose-dependent reduction in caspase 3/7 activity, with the 10 µM dose significantly lowering apoptosis compared to sorafenib-only treatment.

Article Snippet: The caspase 3/7 dye was obtained from Sartorius (Bohemia, NY, USA).

Techniques: Activity Assay, Marker, Activation Assay, Control