Increased M2 Isoform of Pyruvate Kinase in Fibroblasts Contributes to the Growth, Aggressiveness, and Osteoclastogenesis of Odontogenic Keratocysts

Abstract

To investigate the role of glycolysis and the M2 isoform of pyruvate kinase (PKM2) in odontogenic keratocysts (OKCs), the glycolytic flux of primary odontogenic keratocyst fibroblasts (OKC-Fs) and normal oral mucosa fibroblasts (OM-Fs) was determined by glucose uptake, lactate production, and cell proliferation assays. Wound healing assay and Matrigel-coated chamber system were used to investigate the effects of PKM2 on migration and invasion capacities of OKC-Fs. Co-culture of OKC-Fs with osteoclast precursors (RAW264.7 cells) was used to clarify the role of glycolysis in the osteoclastogenic effects of OKC-Fs. In addition, hypoxia-inducible factor 1α and some key enzymes related to glycolysis, including PKM2, 6-phosphofructo-2-kinase/fructose-2, 6-biphosphatase 3, hexokinase 2, and lactate dehydrogenase A, were detected to assess the activation of glycolysis in OKC stroma by immunohistochemistry. Results showed that the glucose uptake and lactate production were significantly higher in OKC-Fs than OM-Fs. PKM2 was elevated in OKC-Fs compared with that in OM-Fs. PKM2 significantly regulated glycolysis, proliferation, migration, invasion, and osteoclastogenic effects of OKC-Fs. Additionally hypoxia-inducible factor 1α, 6-phosphofructo-2-kinase/fructose-2, 6-biphosphatase 3, hexokinase 2, and lactate dehydrogenase A were markedly overexpressed in OKC stroma, and correlated with PKM2. Moreover, the expression of PKM2 was regulated by oxygen concentration in vitro. In sum, PKM2-mediated glycolysis regulated the growth, aggressiveness, and osteoclastogenesis of OKC.