Human Papillary Thyroid Cancer Cells Research Articles

Vernodalin Triggers ROS-Mediated Apoptosis in TPC-1 Human PapillaryThyroid Cancer Cells via Suppression of the MAPKs Signaling Pathway.

Thyroid Cancer (TC) is an endocrine organ malignancy that has become more common in recent decades. Vernodalin (VN), a cytotoxic sesquiterpene, has been reported to exhibit anticancer properties against human breast and liver cancer cells. However, no study has explored the efficacy of VN with respect to its antiproliferative and apoptotic action on human Papillary Thyroid Cancer cells (PTC). The study intended to examine the antitumor and antiproliferative effects of VN and the apoptosis mechanisms underlying its action on TPC-1 human PTC cells. In this study, we examined the VN cell viability by MTT assay; performed ROS measurement by DCFH staining method, MMP identification by Rh-123 staining method, and apoptotic morphological assay by employing AO/EB and DAPI stain method, and further, p38 MAPK/ERK/JNK cell proliferation markers were determined by western blotting technique. The findings showed that VN could inhibit the growth of PTC cells by increasing intracellular ROS, damaging MMP, and stimulating apoptosis in a concentration-dependent manner. The study demonstrated how VN inhibited TPC-1 cell viability by causing ROS-induced cell death via the MAPK signaling pathway. VN may serve as an agonist to impact apoptosis in PTC cells. In human PTC, VN could play an effective role in chemotherapy. More studies pertaining to animal tumor models are needed to prove its anti-cancer effectiveness in vivo.

The common pathogenesis of nodular goiter in both sexes: An exploration into gene expression and signaling pathways

The past few years have witnessed an increasing incidence of nodular goiter (NG), with a well-documented higher prevalence in females than males. This gender disparity has led research to focus primarily on female subjects, potentially overlooking common pathogenic mechanisms in both sexes. In this study, we investigated the shared pathogenesis of NG in males and females. Utilizing a rat model and RNA sequencing, we identified differentially expressed genes associated with the disease. We further validated these findings in normal human thyroid cells and human papillary thyroid cancer cells. A randomized experiment was conducted with equal numbers of male and female rats divided into control and NG model groups. The NG model was established using propylthiouracil and various assessments such as thyroid ultrasonography, thyroid index, thyroid function, and thyroid histology were performed. Transcriptome analysis revealed numerous upregulated and downregulated genes in both male and female model groups. Key genes like KDR, FLT1, PDGFB, and CAV1, and pathways including PI3K-Akt, MAPK, Ras, fluid shear stress and atherosclerosis, calcium signaling, and Rap1 signaling pathways were linked with the disease. Western blot and immunofluorescence analysis confirmed these findings, which were further supported by cell-based experiments. In conclusion, our findings suggest that abnormal expression of specific genes and pathways leading to irregular cell growth, blood vessel formation, and inflammation may be common factors in the pathogenesis of NG in both males and females.

Bergapten attenuates human papillary thyroid cancer cell proliferation by triggering apoptosis and the GSK-3β, P13K and AKT pathways.

Over the past few decades, thyroid cancer (TC) incidence has steadily increased globally. The most common TC is human papillary thyroid carcinoma (PTC), which is poorly responsive to the current treatments. Hence, finding a successful therapeutic is urgently required. Bergapten (BG) is a furanocoumarin, a natural psoralen derivative isolated from numerous species of citrus and bergamot oil that has demonstrated anti-tumor activity. However, there are no reports available on the efficacy of BG on PTC cells. The current research investigated the anti-cancer activity of BG on human BCPAP cells, with cytotoxicity and apoptosis evaluated using MTT assay, AO/EB, DAPI, PI, ELISA, mRNA, and western blot. Bergapten (control group, 10 μM/mL and 15 μM/mL) inhibited PTC cell proliferation and stimulated apoptosis by enhancing Bax and caspase and reducing Bcl-2, cyclin-D1, c-myc, and survivin in a dose-dependent manner. Furthermore, BG expressively attenuated PI3K/AKT/GSK-3β signaling, creating an uneven Bax/Bcl-2 ratio that triggered Cyt-c, caspase cascade and apoptosis in human PTC cells. Our findings emphasize that BG has the potential to be used as a protective natural remedy for human PTC cells.

Adiponectin Inhibits the Progression of Obesity-Associated Papillary Thyroid Carcinoma Through Autophagy.

Obesity is a risk factor for the development of papillary thyroid cancer (PTC). However, the molecular mechanisms by which obesity promotes PTC are unclear. This study aims to identify adipokines that are linked to PTC progression. An adipokine antibody array was used to determine the serum levels of 40 adipokines in normal-weight and obese PTC patients. Enzyme-linked immunosorbent assay was used to determine the serum levels of adiponectin. Recombinant human adiponectin was produced by human adipose-derived stem cells and used to treat PTC cells. Cell proliferation and migration were evaluated using the CCK8 and Transwell assays. Bioinformatics analysis was used to predict mechanisms by which adiponectin affects PTC. Adipokines differentially expressed between normal-weight and obese patients showed a gender-dependent pattern. Obese PTC patients had a significantly lower serum adiponectin level than normal-weight patients, especially in female individuals. Adiponectin levels were negatively correlated with aggressive features of PTC, including tumor diameter > 1 cm, extrathyroidal extension, and lymph node metastasis. Recombinant human adiponectin inhibited the proliferation and migration of human PTC cells in vitro. Bioinformatics analysis identified adiponectin receptor 2 (ADIPOR2) and the autophagy pathway as possible mediators of adiponectin function in TC. In vitro experiments confirmed that adiponectin activated autophagy in PTC cells. These findings shed new lights into the role and mechanisms of adiponectin in TC pathogenesis. Adiponectin is involved in development of obesity-related PTC. Adiponectin can directly inhibit thyroid cancer growth and metastasis through the autophagy pathway.

Identification of Germline FOXE1 and Somatic MAPK Pathway Gene Alterations in Patients with Malignant Struma Ovarii, Cleft Palate and Thyroid Cancer.

Germline variants in the FOXE1 transcription factor have been associated with thyroid ectopy, cleft palate (CP) and thyroid cancer (TC). Here, we aimed to clarify the role of FOXE1 in Portuguese families (F1 and F2) with members diagnosed with malignant struma ovarii (MSO), an ovarian teratoma with ectopic malignant thyroid tissue, papillary TC (PTC) and CP. Two rare germline heterozygous variants in the FOXE1 promoter were identified: F1) c.-522G>C, in the proband (MSO) and her mother (asymptomatic); F2) c.9C>T, in the proband (PTC), her sister and her mother (CP). Functional studies using rat normal thyroid (PCCL3) and human PTC (TPC-1) cells revealed that c.9C>T decreased FOXE1 promoter transcriptional activity in both cell models, while c.-522G>C led to opposing activities in the two models, when compared to the wild type. Immunohistochemistry and RT-qPCR analyses of patients' thyroid tumours revealed lower FOXE1 expression compared to adjacent normal and hyperplastic thyroid tissues. The patient with MSO also harboured a novel germline AXIN1 variant, presenting a loss of heterozygosity in its benign and malignant teratoma tissues and observable β-catenin cytoplasmic accumulation. The sequencing of the F1 (MSO) and F2 (PTC) probands' tumours unveiled somatic BRAF and HRAS variants, respectively. Germline FOXE1 and AXIN1 variants might have a role in thyroid ectopy and cleft palate, which, together with MAPK pathway activation, may contribute to tumours' malignant transformation.

Selenium influences thyroid hormones, myricetin has anti-cancer effects on human papillary thyroid cancer cells, and thyroid hormones have been associated with psychological disorders: Thyroid evaluation and therapy during pregnancy

Introduction: A micronutrient called selenium; a non-metallic element is necessary to manufacture selenoproteins, including selenocysteine. The thyroid has the highest concentration of selenium per gram of tissue in people. The majority of known selenoproteins, including glutathione peroxidase, are expressed in the thyroid and are crucial for the metabolism of thyroid hormones, control of redox states, and preservation of cellular homeostasis. It has been demonstrated that selenium therapy for Graves orbitopathy patients can slow the disease's course and enhance the quality of life. Different selenium supplements have demonstrated varying anticancer efficacy in thyroid cancer. It also looked at how different treatment plans affected patients' thyroid function, mood, and inflammatory markers. TSH (thyroid-stimulating hormone) is slightly elevated in subclinical hypothyroidism, while T4 or T3 levels are normal. Objective: The purpose of this study was to evaluate the therapeutic effects of levothyroxine sodium given alone or in combination with selenium on patients with chronic lymphocytic thyroiditis and hypothyroidism to serve as a guide for clinical treatment. The purpose of the current study is to ascertain the impact of selenium supplementation on thyroid hormone and anti-thyroid peroxidase antibody (anti-TPO AB) levels. The pathophysiology of this illness may be affected by the imbalance in the activity of T cells such as Th1, Th2, Th17, and Treg. This study was to evaluate any changes in the release of cytokines by these cells after selenium administration in patients with HT. The diagnosis and management of thyroid problems during pregnancy are covered in detail in this article. It also includes a summary of the main recommendations made by several medical associations for the diagnosis, treatment, and interpretation of thyroid function during pregnancy. Results: The combined group's overall effective rate was much greater than the control group's. After therapy, both groups' TT3, TT4, TSH, TgAb, and TPOAb levels, SAS and SDS scores, and levels of inflammatory mediators such as IL-2, IL-10, and TNF- all showed a significant improvement. In the combination group, IL-10 levels increased more, and TGAb, TPOAb, IL-2, TNF-, SAS, and SDS scores reduced more when compared to the control group, whereas the differences in the other indices were not statistically significant. According to research, LT4 has some efficacy in treating CLT with hypothyroidism, and selenium therapy in combination with LT4 can increase its therapeutic effect and help patients' immunological and inflammatory responses as well as their mood. Conclusions: Myo-inositol, vitamin D, and selenium supplementation together had an impact on the decline in cytokine markers, Ab-TPO, and Ab-TG levels, which helped to compensate for the underlying condition. Since myricetin is a strong inducer of HATC cell death, it may be helpful in the creation of HATC therapeutic medicines. Myricetin significantly reduced the growth of HATC cells, by almost 70%. A significant number of dead cells showed sub-G1 phase arrest. In SNU-80 HATC cells, myricetin similarly displayed cytotoxicity and caused DNA condensation in a dose-dependent manner. Additionally, myricetin induced the mitochondria to release apoptosis-inducing factor (AIF) into the cytoplasm and changed the potential of the mitochondrial membrane.

Hypoxic papillary thyroid carcinoma cells-secreted exosomes deliver miR-221-3p to normoxic tumor cells to elicit a pro-tumoral effect by regulating the ZFAND5

Papillary thyroid cancer (PTC) has seen a worldwide expansion in incidence in the past three decades. Tumor-derived exosomes have been associated with the metastasis of cancer cells and are present within the local hypoxic tumor microenvironment, where they mediate intercellular communication by transferring molecules including microRNAs (miRNAs) between cells. Although miRNAs have been shown to serve as non-invasive biomarkers for cancer diagnosis, the role of hypoxia-induced tumor-derived exosomes in PTC progression remains unclear. Herein, we investigated the differentially expressed miRNA expression profiles from GEO datasets (GSE191117 and GSE151180) by using the DESeq package in R and identified a novel role for miR-221-3p as an oncogene in PTC development. In vivo and in vitro loss and gain assays were used to clarify the mechanism of hypoxic PTC cells derived exosomal-miR-221-3p in PTC. miR-221-3p was upregulated in human PTC plasma exosomes, tissues and cell lines. We found that hypoxic PTC cells derived exosomal-miR-221-3p promoted normoxic PTC cells proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) in vitro, while inhibition of miR-221-3p limited PTC tumor growth in our PTC xenograft model in nude mice. We finally identified ZFAND5, to be a miR-221-3p target. Mechanistically, hypoxic PTC cell lines-derived exosomes carrying miR-221-3p promoted PTC tumorigenesis by regulating ZFAND5. Our findings further the understanding of the underlying mechanisms associated with PTC progression and identify exosomal-miR-221-3p as a potential biomarker for the diagnosis and prognosis of PTC patients. Our study also suggests that miR-221-3p inhibitors could be a potential treatment strategy for PTC.

Anti-Cancer SERCA Inhibitors Targeting Sorafenib-Resistant Human Papillary Thyroid Carcinoma.

Thyroid cancer is generally curable and, in many cases, can be completely treated, although it can sometimes recur after cancer therapy. Papillary thyroid cancer (PTC) is known as one of the most general subtypes of thyroid cancer, which take up nearly 80% of whole thyroid cancer. However, PTC may develop anti-cancer drug resistance via metastasis or recurrence, making it practically incurable. In this study, we propose a clinical approach that identifies novel candidates based on target identification and validation of numerous survival-involved genes in human sorafenib-sensitive and -resistant PTC. Consequently, we recognized a sarco/endoplasmic reticulum calcium ATPase (SERCA) in human sorafenib-resistant PTC cells. Based on the present results, we detected novel SERCA inhibitor candidates 24 and 31 via virtual screening. These SERCA inhibitors showed remarkable tumor shrinkage in the sorafenib-resistant human PTC xenograft tumor model. These consequences would be clinically worthwhile for the development of a new combinatorial strategy that effectively targets incredibly refractory cancer cells, such as cancer stem cells and anti-cancer drug-resistant cells.

CircRTN1 stimulates HMGB1 to regulate the malignant progression of papillary thyroid cancer by sponging miR-101-3p.

The important role played by circular RNA (circRNA) in promoting the progression of papillary thyroid cancer (PTC) is attracting ever more attention among medical researchers. However, what the precise contribution is of circRTN1 in PTC progression remains unclear. The study was designed to analyze the role and mechanism of circRTN1 in regulating PTC progression. Human PTC cell lines (TPC-1 and IHH-4) and human thyroid normal cells (Nthy-ori 3-1) were used for in vitro assays. mRNA or protein expression of circRTN1, miR-101-3p, and high mobility group box 1 (HMGB1) were detected by quantitative real-time polymerase chain reaction or western blot. Cell proliferation was investigated by cell counting kit-8 assay, cell colony formation assay, and 5-ethynyl-2'-deoxyuridine assay. Wound-healing assay and transwell invasion assay were conducted to evaluate cell migration and invasion. Dual-luciferase reporter assay and RNA immunoprecipitation assay were applied to verify the target relations between circRTN1, miR-101-3p, and HMGB1. A xenograft tumor model was established to demonstrate the effect of circRTN1 on tumor formation in vivo. An immunohistochemistry assay was used to detect protein expression of HMGB1, ki-67, E-cadherin, and vimentin. In comparison with healthy thyroid tissues and cells, PTC tissues and cells displayed high circRTN1 RNA expression and high HMGB1 mRNA and protein expression but low miR-101-3p expression. Silencing of circRTN1 suppressed PTC cell proliferation, migration, and invasion in vitro. MiR-101-3p was a target of circRTN1, and the knockdown of miR-101-3p relieved circRTN1 absence-mediated suppressive effects on PTC cell malignancy. HMGB1 was identified as a target gene of miR-101-3p, and overexpressed HMGB1 almost reverted the inhibitory impacts induced by miR-101-3p mimic in PTC cells. Moreover, circRTN1 silencing hampered tumor formation in vivo. CircRTN1 depletion impeded PTC cell malignancy via the miR-101-3p/HMGB1 pathway, which provided a possible circRNA-targeted therapeutic strategy for PTC.

KMT2D Regulates the NCOA6/THRB Signal Axis through Epigenetic Modification to Promote the Migration and Invasion of Papillary Thyroid Cancer.

The histone lysine methyltransferase Histone-lysine N-methytransferase 2D (KMT2D) is a common mutated gene in a variety of cancers, including papillary thyroid cancer (PTC). However, the mechanism of KMT2D on the progression of PTC remains unclear. In this study, quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting were used to evaluate KMT2D expression between human normal cell (Nthy-ori 3-1) and PTC cells (TPC1, IHH-4 and BCPAP). Proliferation, migration and invasion of TPC1, IHH-4 and BCPAP were assessed by Cell Counting Kit-8 (CCK-8), Wound-healing assay and Transwell assay. The mechanism of KMT2D on thyroid papillary cancer was explored with Chromatin immunoprecipitation assay (ChIP), qRT-PCR and Western blotting. The expression of KMT2D in PTC cells was significantly increased. Downregulation of KMT2D significantly decreased the proliferation, migration and invasion of PTC cells, which was correlated with decreased expression levels of H3K4me2, H3K9me2, NCOA6 and THRB. Meanwhile, ChIP assay demonstrated that KMT2D was associated with NCOA6. Study have shown that the downregulation of KMT2D reduces proliferation, migration and invasion of thyroid papillary carcinoma cells through epigenetic modification of NCOA6/THRB signal axis. These results provide a new insight into the role of KMT2D in migration and invasion of PTC.

Expression of miR-31-5p affects growth, migration and invasiveness of papillary thyroid cancer cells.

In this study, we evaluated the biological role of miRNA-31-5p in papillary thyroid cancer (PTC). By using the real-time PCR, we measured miRNA-31-5p expression levels in 25 PTC tissues and in two human PTC cell lines (K1 and TPC-1). Then, K1 cells were transiently transfected with mirVana inhibitor or mirVana mimic to miRNA-31-5-p. Cell proliferation was determined by MTT and colony formation assays. The in vitro metastatic ability of thyroid cancer cells was evaluated by adhesion, migration and invasion assays. Epithelial mesenchymal transition (EMT) and Hippo pathway related gene and protein levels were evaluated by using the TaqMan™ Gene Expression Assays and western blot analysis, respectively. We found a significant increase of miR-31-5-p expression in tumor tissue and in K1 cells harboring the BRAF p.V600E mutation. Knockdown of miR-31-5p determined a reduction of cell proliferation, associated with a significant decrease in cell adhesion, migration and invasion properties. A downregulation of EMT markers and YAP/β-catenin axis was also observed. Our findings suggest that miRNA-31-5p acts as oncogenic miRNA in human thyrocytes and its overexpression may be involved in the BRAF-related tumorigenesis in PTCs, providing new understanding into its pathological role in PTC progression and invasiveness.

AXL, along with PROS1, is overexpressed in papillary thyroid carcinoma and regulates its biological behaviour

BackgroundAXL, a TAM tyrosine kinase receptor, plays an essential role in the pathogenesis of various solid tumours. This study explores the role of AXL and its ligand PROS1 in the generation and biological behaviour of papillary thyroid cancer (PTC).MethodsThe expression levels of AXL in PTC cancer tissue were analysed using immunohistochemistry (IHC) staining. The expression levels of AXL in PTC and normal thyroid cell lines were analysed using real-time quantitative polymerase chain reaction (RT-qPCR). CCK-8 was used to assess the proliferation of the PTC cell line with and without the effect of the AXL inhibitor (R428). Scratching assays played a role in evaluating the cell migration rate.ResultsPROS1 and AXL were expressed in TPC-1, B-CPAP, and Nthy-Ori 3–1 cells at different levels. Expression was significantly higher in PTC cell lines (TPC-1 and B-CPAP) than in the normal thyroid cell line (Nthy-Ori 3–1) (p < 0.05). In addition, AXL expression in PTC tissues was significantly higher than in adjacent normal tissues (p < 0.05). CCK-8 experiments confirmed that R428 suppresses the proliferation of PTC cell lines in a dose-dependent manner, with an increase in concentration from 0.5 to 4 μM, decreasing the inhibitory effect (p < 0.01). In addition, R428 inhibited PTC cell line migration to different degrees in a range of concentrations from 0.5 to 2 μM compared to control cells (p < 0.01).ConclusionPROS1 and its downstream receptor AXL expression were significantly higher in PTC than in normal thyroid cells. AXL expression was also higher in human PTC tissues than in normal thyroid tissues. Inhibiting the PROS1-AXL-mediated TAM signaling pathway via the AXL blocker R428 suppressed the proliferation and migration of human PTC cells, highlighting the role of this cascade in human PTC development and progression.

Low-Dose Oxidant Toxicity and Oxidative Stress in Human Papillary Thyroid Carcinoma Cells K1

Medical gas plasmas are of emerging interest in pre-clinical oncological research. Similar to an array of first-line chemotherapeutics and physics-based therapies already approved for clinical application, plasmas target the tumor redox state by generating a variety of highly reactive species eligible for local tumor treatments. Considering internal tumors with limited accessibility, medical gas plasmas help to enrich liquids with stable, low-dose oxidants ideal for intratumoral injection and lavage. Pre-clinical investigation of such liquids in numerous tumor entities and models in vitro and in vivo provided evidence of their clinical relevance, broadening the range of patients that could benefit from medical gas plasma therapy in the future. Likewise, the application of such liquids might be promising for recurrent BRAF(V600E) papillary thyroid carcinomas, resistant to adjuvant administration of radioiodine. From a redox biology point of view, studying redox-based approaches in thyroid carcinomas is particularly interesting, as they evolve in a highly oxidative environment requiring the capability to cope with large amounts of ROS/RNS. Knowledge on their behavior under different redox conditions is scarce. The present study aimed to clarify resistance, proliferative activity, and the oxidative stress response of human papillary thyroid cancer cells K1 after exposure to plasma-oxidized DMEM (oxDMEM). Cellular responses were also evaluated when treated with different dosages of hydrogen peroxide and the RNS donor sodium nitroprusside (SNP). Our findings outline plasma-oxidized liquids as a promising approach targeting BRAF(V600E) papillary thyroid carcinomas and extend current knowledge on the susceptibility of cells to undergo ROS/RNS-induced cell death.

Butein induces intrinsic pathway of apoptosis, vimentin proteolysis, and inhibition of cancer stem cell population in a human papillary thyroid cancer cell line

Epithelial-mesenchymal transition (EMT) and cancer stem cells (CSCs) play an essential role in metastasis of papillary thyroid cancer (PTC). Further mesenchymal marker vimentin is linked with metastasis and cancer stem cell generation. Hence, inhibition of EMT and effective elimination of CSCs offers a novel target for the development of new therapeutic agents.The present study observed that at lower concentration, butein, a major bioactive chalcone, significantly inhibits NPA (papillary thyroid cancer cell line) cell migration and reduces extracellular acidification rate (ECAR) an indicator of enhanced glycolysis, required for cell migration. Additionally, at lower concentrations, butein treatment also suppresses vimentin phosphorylation, an essential step in cell migration, proving its potential against cell migration. Phosphorylation of vimentin is crucial in the protection of vimentin from caspase-mediated proteolysis. Interestingly, butein activates caspase-3 for the apoptosis execution at higher concentration; hence, total levels of vimentin were investigated. Butein induces caspase-3 mediated proteolysis of vimentin. Vimentin and glycolysis are essential for maintaining CSCs; therefore, aldeflour assay and side population assay were performed to investigate the effect of butein on CSCs. Our data suggest butein mediates the reduction in CSCs population.Here we report a novel mechanism of butein mediated inhibition of NPA cells migration by suppressing vimentin phosphorylation and its subsequent proteolysis. Collectively our data suggest the potential of butein as an innovative anticancer therapeutic agent for PTC management.

Long non-coding RNA GHET1 promotes thyroid cancer cell proliferation and invasion.

BackgroundWe aimed to evaluate the role of long non-coding RNA (LncRNA) gastric carcinoma proliferation-enhancing transcript 1 (GHET1) on thyroid cancer (TC) behavior in vitro.MethodsTC tissues and paired adjacent normal tissues were obtained after surgical excision from 43 patients with TC. The expression of LncRNA GHET1 was analyzed by real-time (RT) PCR. Human papillary thyroid cancer cell lines (TPC-1, BCPAP) were used to examine the role of LncRNA GHET1 in vitro. Cell proliferation was determined by CCK8 and cell colony formation assays. Transwell and wound-healing assays were used to detect the invasion and migration of thyroid cancer cells.ResultsOur results showed that LncRNA GHET1 was significantly more upregulated in TC tissues than in adjacent normal tissues. LncRNA GHET1 was also increased in thyroid cancer cell lines compared to normal thyroid cell lines. The upregulation of LncRNA GHET1 was significantly associated with tumor invasion, gender, and lymph node metastasis in patients with thyroid cancer. The in vitro studies showed that silencing LncRNA GHET1 in BCPAP cells inhibited cell proliferation, cell invasion, and migration. Silencing of LncRNA GHETI also promoted the cell apoptotic rate, caused an increase in the cell population at the G0/G1 phase, and decreased the cell population at the S phase. In contrast, the overexpression of LncRNA GHET1 promoted cell proliferation, invasion, and migration, inhibited cell apoptosis, and increased cell population at the S phase in TPC cells.ConclusionsLncRNA GHET1 dysregulation might be involved in the carcinogenesis of thyroid cancer. LncRNA GHET1 could be used as a potential molecular marker and molecular target for TC.

Quercetin induces proteolysis of mesenchymal marker vimentin through activation of caspase-3, and decreases cancer stem cell population in human papillary thyroid cancer cell line.

Back groundEpithelial-mesenchymal transition (EMT) and cancer stem cells (CSCs) play an essential role in metastasis of papillary thyroid cancer (PTC), and vimentin is associated with lymph node metastasis and poor prognosis. Hence, inhibition of EMT and effective elimination of CSCs offers a novel target for developing new therapeutic agents. The objective of this study is to inspect the effect of quercetin, a major dietary bioactive flavonoid, on vimentin, CSCs and associated mechanism. MethodsThe first mechanism of quercetin mediated cytotoxicity was investigated in NPA cell line. Subsequently, the effect of quercetin was investigated on glycolysis, vimentin, CSCs, and the mechanism of vimentin degradation was also explored. ResultQuercetin effectively induces cytotoxicity by executing intrinsic apoptosis and activates caspase-3 through extracellular signal-regulated kinase (ERK) activation in NPA cell line having typical mesenchymal characters. Quercetin inhibits NPA cells migration and reduces extracellular acidification rate (ECAR), an indicator of enhanced glycolysis, required for cell migration. Quercetin treatment suppresses vimentin phosphorylation at lower concentrations, a critical step in cell migration, further proving its potential against cell migration. Phosphorylation of Vimentin is also crucial in the protection of vimentin from caspase-mediated proteolysis. Interestingly, quercetin activates caspase-3 at higher concentrations; hence, total vimentin levels were investigated, and caspase-3 mediated proteolysis of vimentin was observed. Vimentin and glycolysis are essential features of the mesenchymal state, which is required to maintain CSCs. Ours data also suggests quercetin mediated reduction in CSCs population. ConclusionWe report a novel mechanism of quercetin mediated inhibition of NPA cells migration by suppressing vimentin phosphorylation and subsequent proteolysis. Collectively our data suggest the potential of quercetin as an innovative anti-cancer therapeutic agent for PTC management.

Taking Advantage of the TGFB1 Biology in Differentiated Thyroid Cancer to Stimulate Sodium Iodide Symporter (NIS)-Mediated Iodide Uptake in Engineered Mesenchymal Stem Cells

The sodium iodide symporter (NIS) mediates the active transport of iodide into thyroid follicular cells, providing the basis for the use of radioiodide for diagnostic imaging and therapy of differentiated thyroid cancer and also non-thyroidal tumors after tumor-selective NIS gene transfer. Based on their excellent tumor-homing capacity, mesenchymal stem cells (MSCs) can be employed as tumor-selective NIS gene delivery vehicles. Transgenic expression of NIS in genetically engineered MSCs allows noninvasive imaging of functional NIS expression as well as therapeutic application of 131I. The use of promoters activated by tumor micromilieu-derived signals to drive NIS expression enhances selectivity and effectiveness, while limiting potential off-target effects. In this study we aimed to exploit the central role of transforming growth factor B1 (TGFB1) in tumor milieu-associated signaling using a TGFB1-inducible synthetic SMAD-responsive promoter to selectively drive NIS-transgene expression in engineered MSCs (SMAD-NIS-MSC) in the context of differentiated thyroid cancer based on the critical role of TGFB1 in the pathogenesis of radioiodine refractory differentiated thyroid cancer. To evaluate the TGFB1 expression in thyroid cancer cell lines, the TGFB1 concentration in conditioned medium (CM) from an array of established human papillary thyroid cancer (PTC) cell lines (BCPAP and K1) was measured by ELISA. BCPAP-CM showed a higher concentration of TGFB1, while a lower concentration was measured in K1-CM. Stimulation of SMAD-NIS-MSCs with PTC-CM showed a significant increase of NIS-mediated radioiodide-125 uptake in these MSCs in vitro. In addition, iodide uptake in SMAD-NIS-MSCs was significantly stimulated by co-culture with thyroid cancer cells. Cell migration assay was performed to validate the effect of PTC-CM in MSC recruitment. MSCs subjected to a gradient between tumor CM and serum free medium showed a directed chemotaxis towards CM with increased forward migration index (FMI) and center of mass (CoM). In a next step, based on the in vitro studies, SMAD-NIS-MSCs will be systemically applied via the tail vein to mice harboring subcutaneous PTC tumors and tumoral iodide uptake will be monitored by 123I-scintigraphy. Taken together, these data indicate the feasibility of commandeering TGF-β/SMAD signaling in the TGFB1-rich tumor environments of radioiodine refractory differentiated thyroid carcinomas to re-establish functional NIS expression using engineered mesenchymal stem cells as therapy vehicles.

SQSTM1/p62 Promotes Cell Growth and Triggers Autophagy in Papillary Thyroid Cancer by Regulating the AKT/AMPK/mTOR Signaling Pathway.

BackgroundThyroid cancer is one of the most common endocrine malignancies worldwide, and papillary thyroid cancer (PTC) is the most common pathologic type of thyroid cancer. SQSTM1/p62 activity mediates different biological functions. This study aimed to investigate the effect of SQSTM1/p62, a multifunctional receptor, on biological function and autophagy characteristics in the human PTC cell line TPC-1.MethodsA total of 105 primary PTC samples and matched adjacent normal thyroid tissue samples were obtained to evaluate the expression of p62 in clinical patients. A similar p62 expression pattern was found in PTC cell lines and normal human thyroid follicular epithelial cells. To evaluate the effect of SQSTM1/p62 on TPC-1 cells, we constructed the p62 knockout cell line p62-KO-TPC-1. Cell proliferation, cell cycle, and cell apoptosis were analyzed by colony formation tests, Cell Counting Kit-8 (CCK-8) assays and flow cytometry in vitro. TPC-1 and p62-KO-TPC-1 human PTC cell lines in the logarithmic growth phase were subcutaneously implanted into BALB/c nude mice to verify their proliferation effect in vivo. Furthermore, western blotting and immunohistochemistry (IHC) were used to detect the expression of AKT/AMPK/mTOR signaling pathway-related proteins.ResultsOverall, p62 expression was higher in tumor tissues than in normal tissues in 73 of 105 PTC patients (69.5%). The expression level of p62 in the PTC cell line was higher than that in the normal thyroid cell line. Our data indicated that in vitro, p62 deficiency could decrease the number of colonies, inhibit cell growth and the cell cycle, and induce apoptosis. Tumor xenograft experiments in BALB/c nude mice corroborated these findings. Moreover, the molecular mechanism was explored by western blotting, and we found that the AMPK/AKT/mTOR pathway was involved.ConclusionsThe results indicate that p62 might mediate cell autophagy and apoptosis in TPC-1 cells via the AMPK/AKT/mTOR pathway and could be used as a potential therapeutic approach for PTC.

In vitro antitumoral effects of the steroid ouabain on human thyroid papillary carcinoma cell lines.

Ouabain is a steroid described as a compound extracted from plants that is capable of binding to Na+ , K+ -ATPase, inhibiting ion transport and triggering cell signaling pathways. Due to its positive ionotropic effect, ouabain was used for more than 200 years for the treatment of cardiac dysfunctions. Numerous antitumor effects of ouabain have been described so far; however, its role on thyroid cancer is still poorly understood. Therefore, the aim of the present work was to evaluate the effect of ouabain on the biology of human papillary thyroid cancer cells. For this, three human thyroid cell lines were used: NTHY-ori, a non-tumor lineage, BCPAP and TPC-1, both derived from papillary carcinomas. Cells were cultured in the presence or absence of ouabain. Subsequently, we evaluated its effects on the viability, cell death, cell cycle, and migratory ability of these cell lines. We also investigated the impact of ouabain in IL-6/IL-6R and epithelial to mesenchymal transition markers expression. Our results indicate that ouabain (10-7 M), decreased the number of NTHY-ori, TPC-1 and BCPAP viable cells and induced cell cycle arrest after in vitro culture, but did not appear to promote cell death. In TPC-1 cells ouabain also inhibited cell migration; increased IL-6/IL-6R expression and IL-6 secretion; and diminished vimentin and SNAIL-1 expression. Collectively, our results indicate that ouabain has an antitumoral role on human papillary thyroid carcinomas in vitro. Even though additional studies are necessary, our work contributes to the discussion of the possibility of new clinical trials of ouabain.

Long noncoding RNA CASC7 inhibits the proliferation and migration of papillary thyroid cancer cells by inhibiting miR-34a-5p

Long noncoding RNAs (lncRNAs) play an essential role in the progression of papillary thyroid cancer (PTC). However, the expression and function of lncRNA cancer susceptibility candidate 7 (CASC7) in PTC remain unknown. The purpose of this study was to investigate the role and molecular mechanism of CASC7 in regulating PTC cell behavior. The expression of CASC7, miR-34a-5p, and tumor protein P73 (TP73) was determined by qRT-PCR and western blot. Cell proliferation was examined by MTT assay. Cell apoptosis was assessed by flow cytometry following Annexin V and PI staining. Cell migration was determined by Transwell migration assay. The interaction between miR-34a-5p and CASC7 or TP73 was examined by luciferase reporter assay. CASC7 and TP73 expression were significantly lower, whereas miR-34a-5p expression was higher in PTC tissues than the adjacent normal tissues. Furthermore, CASC7 overexpression inhibited cell proliferation and migration, whereas facilitated cell apoptosis in human PTC cell lines (K1 and TPC-1). Mechanistically, CASC7 acted as a sponge of miR-34a-5p to upregulate TP73 expression. Moreover, miR-34a-5p mimic transfection could abate the CASC7-regulated PTC cell proliferation, migration, and apoptosis. Collectively, CASC7 inhibited the proliferation and migration of PTC cells by sponging miR-34a-5p to upregulate TP73 expression.