Hemeoxygenase-1 Induces Thyroid Cancer Progression Depending On Subcellular Localization And Enzymatic Activity

Abstract

Thyroid cancer (TC) is the most common endocrine tumor worldwide. Heme Oxygenase-1 (HO-1) is responsible for heme degradation and has shown pro- or antitumor properties depending on the tumor type. In this work, we aimed to study the role of HO-1 in TC biology. For this purpose, we assessed HO-1 expression and localization by Western Blot and indirect immunofluorescence, respectively; cell viability by crystal violet method, cell migration by wound healing assay and cellular protrusions by actin filaments staining. Pharmacological activation of HO-1 with 80μM hemin induced HO-1 overexpression, nuclear localization and increased cell migration and protrusions in TPC-1 (p<0.0001) and 8505c (p<0.0001) cell lines. In the Nthy-Ori- 3-1 non-malignant cell line, only HO-1 cytoplasmic localization was observed and cell migration was unaffected. Pharmacological inhibition of HO-1 with 16μM ZnPP induced HO-1 expression, cytoplasmic localization and reduced migration in TPC-1, 8505c and Nthy-Ori-3-1 (p<0.001) cells. To evaluate nuclear HO-1 contribution in TC cell viability, we treated TPC-1 cells with 10μM E-64 (a cysteine- protease inhibitor shown to inhibit HO-1 nuclear translocation) or its combination with hemin. Both the treatment with E-64 alone and with E-64+hemin showed a reduction in HO-1 nuclear expression. In addition, E-64+hemin showed a decrease in cell viability compared with hemin treatment alone (48-72h, p<0.0001). Since MAPK is the main signaling pathway involved in TC, we analyzed pERK levels and observed that it increased after hemin treatment. Moreover, 80μM hemin, 10μM U0126 (MEK inhibitor) or their combinations, failed to alter HO-1 expression, thus showing that HO-1 is located upstream of the MEK-ERK pathway. In conclusion, our results indicate a protumor role for HO-1 in TC depending on its subcellular localization and enzymatic activity suggesting that both the canonical and non-canonical pathways of HO-1 are involved in TC progression