Abstract
BackgroundTumor invasiveness is directly related to the ability of tumor cells to migrate and invade surrounding tissues, usually degrading extracellular matrix. Despite significant progress in the knowledge about migration and invasion, there is much more to elucidate about their regulatory mechanisms, especially in cancer cells. MicroRNAs (miRs) were recently described as important regulators of migration. Differential expression of miRs in cancer is frequently associated with progression, invasion and metastasis. In papillary thyroid carcinoma (PTC), miR-146b-5p is highly expressed and positively correlated to the degree of malignancy.MethodsThis study aimed to investigate the role of miR-146b-5p on the migratory and invasive behaviors of thyroid cells, using a non tumor rat thyroid follicular cell line (PCCl3) transfected with the miR-146b-5p genomic region, and two PTC cell lines (TPC-1 and BCPAP, bearing distinct oncogenic backgrounds), which express high levels of miR-146b-5p, after miR-146b inhibition by antagomiR and miR-146b overexpression by mimics-miR. Migration and invasion were studied by time-lapse and transwell assays (with and without Matrigel®). Gelatin degradation assays were also employed, as well as F-actin staining.ResultsMigration and invasion of PCCl3 were increased 2-3x after miR-146b-5p overexpression (10X) and large lamellipodia were evident in those cells. After miR-146b-5p inhibition, TPC-1 and BCPAP migration and invasion were significantly reduced, with cells showing several simultaneous processes and low polarity. Gelatin degradation was inhibited in TPC-1 cells after inhibition of miR-146b-5p, but was unaffected in BCPAP cells, which did not degrade gelatin. The inhibition of miR-146b-5p in PCCl3 also inhibited migration and invasion, and additional (exogenous) overexpression of this miR in TPC-1 and BCPAP cells increased migration and invasion, without effects on cell morphology or gelatin degradation. The overexpression of SMAD4 in BCPAP cells, a validated target of miR-146b-5p and key protein in the TGF-β signaling pathway, inhibited migration similarly to the effects observed with the antagomiR 146b-5p.ConclusionsmiR-146b-5p positively regulates migration and invasion of thyroid normal and tumor follicular cells (independently from their original mutation, either BRAF or RET/PTC), through a mechanism that involves the actin cytoskeleton but not an increased capacity of matrix degradation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-016-2146-z) contains supplementary material, which is available to authorized users.
Highlights
Tumor invasiveness is directly related to the ability of tumor cells to migrate and invade surrounding tissues, usually degrading extracellular matrix
We aimed to investigate the influence of miR-146b-5p on the migratory and invasive behaviors of thyroid normal and tumor cells, in order to determine its contribution to thyroid carcinoma invasiveness
The overexpression of SMAD4 in BCPAP cells, a validated target of miR-146b-5p and key protein in the TGF-β signaling pathway, inhibited migration to the effects observed with the antagomiR 146b-5p
Summary
Tumor invasiveness is directly related to the ability of tumor cells to migrate and invade surrounding tissues, usually degrading extracellular matrix. Tumor invasiveness is directly related to the ability of tumor cells to migrate and invade surrounding tissues, spreading via blood and lymphatic circulation. Invasive adherent tumor cells present a mesenchymal phenotype and are able to migrate faster, degrading extracellular matrix on their way. Sometimes filopodia, which are thin spike-like exploratory processes, precede or accompany lamellipodia formation. This migration cycle is regulated by Rho GTPases, central modulators of the cytoskeleton involved in many signaling pathways [4]. The classic regulatory cycle of Rho GTPases involve molecules that regulate GTP binding and hydrolysis, as well as the availability of GTPases to be activated, usually in cell membranes. In the last few years, other important regulatory mechanisms were described, including microRNAs (miRs) [5]
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