Decreased Liver Tumorigenesis in Sulfatase 2 (SULF2) Knockout Mice May Be Mediated Through Inhibition of the Epithelial-Mesenchymal Transition (EMT)

Abstract

Background and Aim. The pathogenesis and progression of pancreatic cancer involves conversion of TGFβ suppressive signaling in early lesions to an unmasked, TGFβ proliferative signaling cascade in metastatic lesions. We have previously demonstrated that TGFβ proliferative signaling downregulates the tumor suppressor PTEN and induces motility of pancreatic cancer cells after ligand-receptor binding with subsequent increase in cytoplasmic free Ca2+ concentration ([Ca2+]cyt) and activation of PKCα and NF-κB. [Ca2+]cyt homeostasis is important to maintain cell functionality dysregulation of which contributes to tumorigenesis of pancreas. Transient receptor potential ion channels (TRPC) help regulate [Ca2+]cyt, and here we evaluated the role of TRPC channels in regulating TGFβ proliferative signaling. Methods. BxPc3 pancreatic cancer cells were treated with TGFβ (10 ng/mL), and TRPC and PTEN expression were analyzed by RT-PCR and/or Western blots. [Ca2+]cyt was assessed using Fura-2 fluorescence ratio digital imaging. Pharmacological reagents (TRPC inhibitors: 2-APB, SKF96365, and LaCl3) and siRNAs to TRPCs were used to determine the role of TRPCs on TGFβ-induced PTEN regulation and cell migration. Cell motility was assessed by Boyden chamber migration assay. Results. BxPc3 cells express TRPC1, 4, and 6 isoforms. TGFβ induced an increase in [Ca2+]cyt in Ca2+-free solutions and also induced an increase in [Ca2+]cyt when extracellular Ca2+ was restored. TGFβ-induced Ca2+ entry involves the activation of TRPC, as the TRPC inhibitors 2-APB, SKF96365, and LaCl3, were able to reduce peak [Ca2+]cyt by 80%, 40%, and 29%, respectively, and reversed TGFβ-induced PTEN downregulation. The 2-APB, SKF96365, and LaCl3 inhibitors also reduced TGFβinduced cell motility. Using siRNA, inhibition of TRPC1 but not other isoforms, successfully prevented TGFβ-induced cell motility. Conclusion. TRPC1 regulates TGFβ-induced downregulation of PTEN and pancreatic cell motility. This aspect of TGFβ regulation of PTEN and pancreatic cell motility might be exploited therapeutically to control pancreatic cancer metastasis.