KLF2 Promotes Ellagic Acid-Mediated Osteogenic Differentiation of Dental Pulp-Derived Stem Cells via Autophagy and Mitochondrial Regulation

Backgrounds: Krüppel-like factor 2 (KLF2), a zinc-finger transcription factor of the Kruppel-like factor family, plays a crucial role in regulating various cellular processes, including differentiation, autophagy, and metabolism. However, it is not clear whether it has any role in the ellagic acid (EA)-mediated osteoblastic differentiation of dental pulp-derived stem cells (DPSCs). To investigate any regulatory role of KLF2 during EA-induced osteoblastic differentiation of DPSC, we have evaluated the path-ways emphasizing autophagy, mitophagy, and mitochondrial bioenergetics. Methods: We used induction and reduction of KLF2 approaches using chemical compounds, such as geranylgeranyl transferase inhibitor 298 (GGTI298), a known inducer of KLF2, and geranylgeranyl pyrophosphate (GGPP), a known inhibitor of KLF2. The key osteogenic, autophagy, and mitophagy markers were assessed via RT-qPCR and Western blotting, intracellular and mitochondrial ROS, along with mitochondrial membrane potential using high-resolution confocal microscopy, and cellular bioenergetics using Seahorse XF methods. Results: We found that EA alone significantly upregulated osteogenic markers, along with enhanced expression of autophagy and mitophagy-related molecules, and mitochondrial biogenetics. However, when we induced the KLF2, it amplified the expression of these markers and improved mitochondrial bioenergetics, suggesting a distinct relation between EA and KLF2. On the other hand, inhibition of KLF2 led to a significant downregulation of these markers. Conclusion: This supports the notion that KLF2 is a pivotal transcriptional regulator involved in mediating the pro-osteogenic effects of EA. It's activation enhanced autophagy, and improved mitochondrial bioenergetics, thereby facilitating EA-induced osteogenic differentiation of DPSCs. These findings refine our understanding of molecular mechanisms underlying polyphenol-mediated osteogenesis and show the pivotal role of KLF2 for regeneration via osteogenic differentiation of DPSCs.