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 pathways 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.
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