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Peroxisome proliferator-activated receptors (PPARs) are a class of ligand-activated nuclear receptor transcription factors belonging to the nuclear receptor superfamily subfamily 1C (NR1C). The PPAR family comprises three highly homologous isoforms: PPARα (NR1C1), PPARδ (also known as PPARβ; NR1C2), and PPARγ (NR1C3). PPARα is highly expressed primarily in the liver, heart, kidney, and brown adipose tissue, serving as a key regulator of fatty acid oxidation and lipid metabolism; PPARγ is abundant in white adipose tissue and immune cells, playing a central role in adipocyte differentiation, insulin sensitization, and the regulation of inflammation; PPARδ exhibits the broadest expression profile, being distributed across tissues such as skeletal muscle, cardiac muscle, liver, adipose tissue, skin, and macrophages. All PPAR isoforms share a classic nuclear receptor structure: a highly conserved DNA-binding domain (DBD) responsible for recognizing peroxisome proliferator response elements (PPREs) within target gene promoters, and a C-terminal ligand-binding domain (LBD) responsible for binding endogenous or exogenous ligands and mediating ligand-dependent transcriptional activation. The PPAR family plays an indispensable role in energy metabolism, cell differentiation, inflammation regulation, and the pathogenesis of metabolic diseases (such as diabetes, obesity, and atherosclerosis) and tumorigenesis.

HEK293 Human PPARδ-GAL4 Luciferase Reporter Cell engineered to express the luciferase reporter gene under the control of PPARδ-GAL4. GW0742 is a potent agonist of PPARβ and PPARδ; The underlying principle is illustrated in the figure below.

Figure 1. Schematic Diagram Illustrating the Principle of the HEK293 Human PPARδ-GAL4 Luciferase Reporter Cell

Figure 2. Dose Response of GW7042 in PPARδ-GAL4 Luciferase Reporter HEK293(C32).

Cell Resuscitation
1)Rapidly thaw the frozen cells in a 37 °C water bath for approximately 60 seconds. Once thawed (which may take slightly less or more than 60 seconds), immediately transfer the cell suspension from the cryovial into a 15 mL centrifuge tube containing 10 mL of pre-warmed HEK293 Human PPARδ-GAL4 Luciferase Reporter Cell complete culture medium.
2)Centrifuge cells at 1000 rpm for 5 min to remove medium, then resuspend cells in 5 mL of pre-warmed complete medium.
3)Transfer the cell suspension into a T25 culture flask and incubate at 37 °C with 5% CO₂.
4)After approximately 24–36 hours, replace the medium or passage the cells to remove non-adherent dead cells.

Subculturing procedure
1)When the cell density reaches the appropriate confluency for passaging, wash the cells with PBS, then add 1 mL trypsin to detach the cells. When more than 80% of the cells detach upon gently tapping the culture flask, add complete culture medium to terminate digestion. Gently pipette to obtain a single-cell suspension, transfer to a 15 mL centrifuge tube, and centrifuge at 1000 rpm for 5 minutes.

2)Discard supernatant after centrifugation. Resuspend cells in fresh medium to a single-cell suspension and transfer to a new culture flask for continued growth.

Cell Freezing
After trypsinization and centrifugation of cells from each T75 flask or 10 cm culture dish, discard the supernatant. Add 2 mL of cryopreservation medium (90% FBS + 10% DMSO), gently resuspend thoroughly, and aliquot into two cryovials. Immediately place the cryovials into a controlled-rate freezing container (e.g., Nalgene 5100-0001), fill with isopropanol to the indicated level, and store at −80 °C. After 24 hours, transfer the cryovials to liquid nitrogen for long-term storage.