My cells are primary human aortic valve cells, treated with ATF3 Inducer 1 (CAS No.: 3038756-30-5). Culture conditions: cultured in DMEM high glucose medium and osteogenic medium (OM) for two weeks, changing the medium every three days, adding ATF3 inducer 1 at a concentration of 50 μM when changing the medium, and performing WB to detect protein levels after two weeks of culture as shown in the attached figure. From left to right are the control cultured in DMEM medium, solvent control and ATF3 inducer 1; the control cultured in OM medium, solvent control (DMSO) and ATF3 inducer 1.
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ATF3 is a typical "stress response gene" that is rapidly upregulated after being subjected to a variety of stimuli (cell stress, damage, proliferation promotion, etc.), but then suppresses its own expression and restores homeostasis through a complex negative feedback mechanism.
The ATF3 Inducer 1, you used is at a high concentration (50μM), and the incubation time is as long as two weeks, with the drug replenished every three days. This condition may cause ATF3 to be activated, and the cells continue to suppress its expression through the above-mentioned multiple negative feedback mechanisms, even below the basal level. It is recommended to use qPCR to detect short-term (1h, 4h, 24h) and long-term (1-2 weeks) changes in ATF3 mRNA to verify whether there is a dynamic process of "first rise and then fall".
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