Supplementary MaterialsAdditional file 1: Figure S1. Error bars represent means standard errors of the mean (SEM) from three independent experiments. B. Cell cycle assay. BcPAP and TPC1 cells after siPDPN or siNeg transfection and control cells (no siRNA) were fixed, permeabilized, stained with propidium iodide (PI) solution, and then analyzed by flow cytometry. The results are presented as FTDCR1B percentage of cells in G1, S, and G2/M phases. (ZIP 907 kb) 12885_2018_5239_MOESM1_ESM.zip (908K) GUID:?6AF9EDBD-2537-420F-98A2-5A69D0A84D0C Data Availability StatementThe datasets used and/or analyzed in this study may be received from the corresponding author on reasonable request. Abstract Background Podoplanin (PDPN) is a mucin-type transmembrane glycoprotein specific to the lymphatic system. PDPN expression has been found in various human tumors and is considered to be a marker of cancer. We had previously shown that PDPN expression contributes to carcinogenesis in the TPC1 papillary thyroid cancer-derived cell line by enhancing cell migration and invasiveness. The aim of this Bafetinib (INNO-406) study was to determine the effect of PDPN down-regulation in another thyroid cancer-derived cell line: BcPAP. Methods In order to determine the effects of PDPN on malignant features of BcPAP cells (harboring the mutated allele) and TPC1 cells (carrying the rearrangement), we silenced PDPN in these cells using small interfering RNA (siRNA). The efficacy of PDPN silencing was confirmed by qRT-PCR and Western blotting. Then, we tested the motility and invasiveness of these cells (using scratch test and Transwell assay), their growth capacities F(cell Bafetinib (INNO-406) cycle analysis, viability, clonogenic activity) and apoptosis assays), adhesion-independent colony-formation capacities, as well as the effect of PDPN silencing on MMPs expression and activity (zymography). Results We found that PDPN-induced cell phenotype depended on the genetic background of thyroid tumor cells. PDPN down-regulation in BcPAP cells was negatively correlated with the migration and invasion, as opposed to TPC1 cells where PDPN depletion led to improved invasiveness and migration. Moreover, our outcomes claim that in BcPAP cells, PDPN could be mixed up in epithelial-mesenchymal changeover (EMT) through regulating the manifestation from the ezrin, radixin and moesin (E/R/M) protein, MMPs 9 and MMP2, redesigning of actin cytoskeleton and mobile protrusions. We also proven that PDPN manifestation is from the MAPK signaling pathway. The inhibition from the MAPK pathway led to a reduced PDPN manifestation, improved E/R/M phosphorylation and decreased cell migration. Additionally, PDPN depleted BcPAP cells treated with inhibitors of MEK1/2 kinases (U0126) or from the BRAF V600E proteins (PLX4720) had decreased motility, much like that seen in TPC1 cells following PDPN knock-down previously. Conclusions Completely, our data claim that PDPN may play a significant role within the control of Bafetinib (INNO-406) invasion and migration of papillary thyroid carcinoma cells in colaboration with the E/R/M, MAPK and MMPs kinases. Electronic supplementary materials The online edition of this content (10.1186/s12885-018-5239-z) contains supplementary materials, which is open to certified users. mutation, possess higher PDPN manifestation level. This might claim that this gain-of-function mutation may be connected with a stronger induction of PDPN expression . Therefore, we prolonged our analyses towards the BcPAP cell range harboring a mutated allele (can be a common mutation that takes on an essential part in tumorigenesis and development of PTC [32C34]. Although signaling pathways triggered by and overlap, the tumors connected with each one of these two modifications have exclusive phenotypic features, recommending that they could possess different tumor biology [35C39] also. Therefore, in today’s study,.