OBJECTIVES: Lymph node metastasis in papillary thyroid cancer (PTC) is closely associated with tumor recurrence and patient survival. However, current technologies have limited sensitivity in detecting occult cervical lymph node metastases. Identifying accurate molecular markers for predicting PTC metastasis holds significant clinical value. This study aims to analyze WNT10A expression in PTC and its clinical significance, and to explore the role of WNT10A gene knockdown in PTC cell proliferation, invasion, and metastasis. METHODS: The expression of WNT10A in thyroid carcinoma was analyzed using the Gene Expression Profiling Interactive Analysis (GEPIA) and University of Alabama at Birminghara Cancer data analysis Portal (UALCAN) databases. Real-time RT-PCR was used to measure WNT10A mRNA levels in tumor and adjacent normal tissues from 32 PTC patients. Immunohistochemistry was conducted on 158 PTC specimens to assess WNT10A protein expression and its correlation with clinicopathological features. In vitro experiments were performed using K1 and TPC-1 cell lines. Cell proliferation was assessed using the Celigo system and methyl thiazolyl tetrazolium (MTT) assays; apoptosis was measured via flow cytometry; invasion and metastasis were evaluated using scratch and Transwell assays. A xenograft model was established in nude mice to observe tumor growth, and tumor weight and volume were compared between cell lines. Differentially expressed genes regulated by WNT10A were identified via mRNA sequencing, followed by Gene Ontology (GO) and ingenuity pathway analysis (IPA). Real-time PCR and Western blotting were used to validate the effects of WNT10A on key downstream mRNA and protein in the Tec kinase signaling pathway. RESULTS: WNT10A mRNA expression was significantly higher in thyroid cancer tissues compared to adjacent normal tissues according to GEPIA and UALCAN (both P<0.01). The real-time RT-PCR result showed that WNT10A mRNA expression in PTC tissues was high than that in adjacent tissues (P<0.01). Immunohistochemistry revealed significantly higher WNT10A protein expression in PTC tissues compared to adjacent tissues (P<0.01), and its expression correlated with multifocality, extrathyroidal invasion, and lymph node metastasis. WNT10A knockdown significantly inhibited proliferation, altered cell cycle distribution, and increased apoptosis in K1 and TPC-1 cells (all P<0.01). WNT10A silencing also reduced migration and invasion abilities in both cell lines. In vivo, WNT10A knockdown in TPC-1 cells suppressed tumor formation in nude mice. GO analysis and IPA suggested that the Tec kinase signaling pathway was a key downstream target of WNT10A. RT-PCR and Western blotting confirmed that WNT10A knockdown downregulated the expression of key genes (STAT3, MAPK8, TNFRSF21, and AKT2) in this pathway. CONCLUSIONS WNT10A is highly expressed in PTC and is associated with tumor proliferation, invasion, and metastasis. Its tumor-promoting effects may be mediated through suppression of the Tec kinase signaling pathway.
Humans ; Cell Proliferation ; Thyroid Cancer, Papillary/pathology* ; Thyroid Neoplasms/metabolism* ; Animals ; Wnt Proteins/metabolism* ; Neoplasm Invasiveness ; Mice ; Cell Line, Tumor ; Female ; Male ; Mice, Nude ; Apoptosis ; Lymphatic Metastasis ; Middle Aged ; Cell Movement ; Adult

OBJECTIVES: Increasing detection of low-risk papillary thyroid carcinoma (PTC) is associated with overdiagnosis and overtreatment. N6-methyladenosine (m⁶A)-mediated microRNA (miRNA) dysregulation plays a critical role in tumor metastasis and progression. However, the functional role of m⁶A-miRNAs in PTC remains unclear. This study aims to elucidate the regulatory mechanism of m⁶A-miR-139-5p expression in PTC, determine its association with PTC metastasis, and evaluate its potential as a diagnostic biomarker for PTC metastasis, thereby providing experimental evidence for precision diagnosis and therapy. METHODS: Expression profiles of m⁶A-miRNAs were compared between the The Cancer Genome Atlas (TCGA) and GSE130512 cohorts to identify metastasis-associated candidates. Clinical specimens from 13 metastasis and 18 non-metastasis PTC patients were analyzed to assess m⁶A-miR-139-5p expression and its correlation with metastasis. Functional experiments were conducted to investigate the effect of fat mass and obesity-associated protein (FTO) on pri-miR-139 methylation and processing, clarifying its regulatory role in miR-139-5p expression. In TPC-1 cells, MTT assays were performed to evaluate whether miR-139-5p overexpression could counteract FTO-mediated cell proliferation. Transwell invasion assays were used to determine the impact of miR-139-5p on PTC cell invasion, exploring whether it functions through the ZEB1/E-cadherin axis. RESULTS: By comparing TCGA and GSE130512 cohorts, it was found that circulating m⁶A-miR-139-5p could serve as a biological indicator for detecting PTC metastasis. Detection of 13 metastatic and 18 non-metastatic clinical specimens showed that FTO inhibited the processing of pri-miR-139 by reducing its methylation level, leading to the dysregulation of miR-139-5p in PTC (P<0.05). In TPC-1 cells, MTT assay showed that overexpression of miR-139-5p could partially reverse FTO overexpression-mediated cell proliferation (P<0.05). In addition, miR-139-5p inhibited the invasive ability of PTC cells by targeting the ZEB1/E-cadherin axis, while FTO overexpression could partially weaken this inhibitory effect. CONCLUSIONS Circulating miR-139-5p can be a potential marker for evaluating PTC metastasis. FTO affects the expression and function of miR-139-5p by regulating m⁶A modification of pri-miR-139, but its clinical value needs further verification.
Humans ; MicroRNAs/metabolism* ; Thyroid Cancer, Papillary/metabolism* ; Alpha-Ketoglutarate-Dependent Dioxygenase FTO/metabolism* ; Thyroid Neoplasms/metabolism* ; Cell Line, Tumor ; Neoplasm Metastasis ; Adenosine/genetics* ; Gene Expression Regulation, Neoplastic ; Female ; Male ; Cadherins/metabolism* ; Cell Proliferation ; Zinc Finger E-box-Binding Homeobox 1/genetics*

OBJECTIVES: To investigate the expression of apolipoprotein C1 (APOC1) in papillary thyroid carcinoma (PTC) and its effects on proliferation and apoptosis of PTC cells. METHODS: The expression level of APOC1 in PTC and its impact on prognosis were analyzed using GEPIA 2 and Kaplan-Meier databases. Immunohistochemistry (IHC) and Western blotting were used to detect the expression of APOC1 in PTC and adjacent tissues and in 3 PTC cell lines and normal thyroid Nthyori 3-1 cells. In TPC-1 and BCPAP cells, the effect of Lipofectamine 2000-mediated transfection with APOC1 siRNA or an APOC1-overexpressing plasmid on cell growth and colony formation ability were examined by observing the growth curves and using colony-forming assay. The changes in cell cycle and apoptosis of the transfected cells were analyzed with flow cytometry. RT-qPCR and Western blotting were used to detect the changes in expressions of P21, P27, CDK4, cyclin D1, Bcl-2, Bax, caspase-3 and caspase-9 and the key proteins in the JAK2/STAT3 signaling pathway. RESULTS: APOC1 expression was significantly higher in PTC tissues and the 3 PTC cell lines than in the adjacent tissues and Nthyori 3-1 cells, respectively. In TPC-1 and BCPAP cells, APOC1 knockdown obviously reduced cell proliferative activity, increased the percentage of G0/G1 phase cells, lowered the percentages of S and G2 phase cells, promoted cell apoptosis, and downregulated mRNA and protein expression levels of CDK4, cyclin D1 and Bcl-2 and the protein levels of p-JAK2 and p-STAT3. APOC1 overexpression in the cells produced the opposite effects on cell proliferation, apoptosis, cell cycle and the mRNA and protein expressions. The application of AG490, a JAK2 inhibitor, strongly attenuated APOC1 overexpression-induced activation of the JAK2/STAT3 signaling pathway in BCPAP cells. CONCLUSIONS APOC1 overexpression promotes proliferation and inhibits apoptosis of PTC cells possibly by activating the JAK2/STAT3 signaling pathway and accelerating cell cycle progression.
Humans ; Apoptosis ; Cell Proliferation ; STAT3 Transcription Factor/metabolism* ; Signal Transduction ; Janus Kinase 2/metabolism* ; Thyroid Neoplasms/pathology* ; Thyroid Cancer, Papillary ; Cell Line, Tumor ; Carcinoma, Papillary

OBJECTIVE: To assess the value of DNA methylation level of HYAL2 gene as a molecular marker for differential diagnosis of malignant and benign thyroid tumors. METHODS: DNA methylation of HYAL2 gene in tissue specimens of 190 patients with papillary thyroid cancer (PTC) and 190 age- and gender-matched patients with benign thyroid tumors was examined by mass spectrometry, and the protein expression of HYAL2 was detected immunohistochemically for another 55 pairs of patients. Logistic regression analysis was performed to calculate the odds ratio (OR) and evaluate the correlation of per 10% reduction in DNA methylation with PTC. Receiver operating characteristic (ROC) curve analysis was performed and the area under curve (AUC) was calculated to assess the predictive value of alterations in HYAL2 methylation. RESULTS: Hypomethylation of HYAL2_CpG_3 was significantly correlated with early-stage PTC (OR=1.51, P=0.001), even in stage I cancer (OR=1.42, P=0.007). Age-stratified analysis revealed a significantly stronger correlation between increased HYAL2_CpG_ 3 methylation and early-stage PTC in patients below 50 years than in those older than 50 years (OR: 1.89 vs 1.37, P < 0.05); ROC analysis also showed a larger AUC of 0.787 in younger patients. The results of immunohistochemistry showed that patients with PTC had significantly higher protein expressions of HYAL2 than patients with benign tumors. CONCLUSION The alterations of DNA methylation level of HYAL2 gene is significantly correlated with early-stage PTC, suggesting the value of DNA methylation level as a potential biomarker for differentiation of malignant from benign thyroid tumors.
Adenoma, Oxyphilic/genetics* ; Biomarkers, Tumor/metabolism* ; Cell Adhesion Molecules/metabolism* ; DNA Methylation ; GPI-Linked Proteins/metabolism* ; Humans ; Hyaluronoglucosaminidase/metabolism* ; Immunohistochemistry ; Middle Aged ; Thyroid Cancer, Papillary/pathology* ; Thyroid Neoplasms/pathology*

OBJECTIVE: To investigate the expression and clinical significance of EphA2 and E cadherin proteins in papillary thyroid carcinoma tissues, and to explore the relationship between them. METHOD: Using immunohistochemical SP/PV method, we detected the expression of EphA2 and E cadherin in tumors of 43 papillary thyroid carcinomas, 11 thyroid adenoma and 10 normal thyroid tissues, then studied their relationships with clinic pathological factors. RESULT: The total positive rates of EphA2 and E cadherin expression were 58. 14% and 32. 56% in papillary thyroid carcinoma tissues, 18. 18% and 81. 81% in thyroid adenoma.tissues and they were 10. 00% and 100. 00% in normal thyroid tissues respectively. The positive expression of EphA2 in carcinoma tissues was higher than in the thyroid adenoma tissues and normal thyroid tissues (P<0. 05) and the positive expression of E cadherin in carcinoma tissues was lower than that in the thyroid adenoma tissues and normal thyroid tissues (P<0. 05). The positive expression of EphA2 and E cadherin was associated with lymph node metastasis and histological grade (P<0. 05), but it was not associated with all the clinic-pathological factors including age, sex and the tumor size (P>0. 05). In papillary thyroid carcinoma tissues, the expression of EphA2 was negatively correlated with the expression of E cadherin protein (r= -0. 416, P<0. 01). CONCLUSION EphA2 and E cadherin may be involved in carcinogenesis and development of papillary thyroid carcinoma.
Adenoma ; metabolism ; pathology ; Antigens, CD ; Cadherins ; metabolism ; Carcinoma ; metabolism ; pathology ; Carcinoma, Papillary ; Humans ; Lymphatic Metastasis ; Receptor, EphA2 ; metabolism ; Thyroid Cancer, Papillary ; Thyroid Gland ; metabolism ; Thyroid Neoplasms ; metabolism ; pathology

OBJECTIVE: To study the expression of trefoil factor 3 (TFF3)with stromal cell-derived factor (SDF-1) and its receptor (CXCR4) in papillary thyroid carcinoma(PTC), and investigate the function of TFF3, SDF-1/ CXCR4 and the relationship among them during the tumor genesis,development and outcome of PTC. METHOD: Detecting the expression of TFF3 and SDF-1/CXCR4 by immunohistochemical method (SP) in 92 cases of PTC and para-carcinoma tissue. Semiquantitative analysis of the results of immunohistochemistry was conducted by image analysis software. RESULT: (1) TFF3 protein was expressed in the cytoplasm of cancer cells,while TFF3 was negative or weakly positive in follicular cells of para-carcinoma tissue. The positive expression rate of TFF3 was 92.39%, of which the strong positive rate of clinical stage III-IV accounted for 71.79% (42/59) and that of clinical stage I-II was 33.33% (11/33) (P < 0.01). The positive rate of TFF3 was significantly higher in the cases with lymph node metastasis than those without lymph node metastasis (100.00% vs 86.27%, P < 0.05). The AOD value of TFF3 was higher in PTC than in para-carcinoma tissue, that in cases with lymph node metastasis was higher than those without lymph node metastasis, and that in stage III-IV was higher than those in I-II (P < 0.05 or P < 0.05). (2) There was high expression of SDF-1 in the cytoplasm of malignant tissues. The para-carcinoma tissue was weakly positive or negative to SDF 1 and metastatic lymph nodes was weakly positive to SDF 1. The positive rates and AOD values of SDF-1 protein were similar to those of TFF3 in PTC,that is to say the positive rate and AOD values were higher in PTC than in para-carcinoma tissue, those in cases with lymph node metastasis were higher than those without lymph node metastasis, those in stage III-IV were higher than those in I-II, and those in patients older than 45 years old was obviously higher than those in patients under 45 years old (P < 0.05 or P < 0.01); CXCR4 was also mainly expressed in cytoplasm with few expression in nuclei, while negative or weakly positive in para-carcinoma. The positive rate and AOD values of CXCR4 in PTC were similar to SDF-1, meaning that they were higher in PTC than in para-carcinoma tissue and associated with the clinical stage, lymph node metastasis and age (P < 0.05 or P < 0.01). (3) There was positive relationship between TFF3 and SDF-1 as well as between SDF-1 and CXCR4 in PTC (r = 0.971, P < 0.01). CONCLUSION The high expression of TFF3, SDF-1 and CXCR4 in PTC are correlated with carcinogenesis and progression, and may play a significant role in evaluating the malignancy degree and progression of PTC.
Adult ; Aged ; Carcinoma ; metabolism ; pathology ; Carcinoma, Papillary ; Chemokine CXCL12 ; metabolism ; Female ; Humans ; Lymphatic Metastasis ; Male ; Middle Aged ; Neoplasm Staging ; Peptides ; metabolism ; Receptors, CXCR4 ; metabolism ; Signal Transduction ; Thyroid Cancer, Papillary ; Thyroid Neoplasms ; metabolism ; pathology ; Trefoil Factor-2 ; Young Adult

OBJECTIVE: To relatively detect the Runx2 mRNA expression in papillary thyroid carcinoma (PTC) and thyroid adenoma, then to investigate the role of Runx2 in the development and progression in PTC and the relationship with the micro calcification in PTC. METHOD: The expression of Runx2 mRNA in 14 samples of PTC and 14 samples of thyroid adenoma was examined by relatively real-time RT-PCR. RESULT: The deltaCT value of the carcinoma group and adenoma group was 2.395 +/- 0.302 and 5.028 +/- 1.179 respectively (P<0.01). The 2(-deltadeltaCT) value of the carcinoma group and adenoma group was 7.826 +/- 5.004 and 1 respectively (P<0.01). The carcinoma group was divided into two groups by calcification and there was no statistical difference (P>0.05), and the adenoma group as well. The carcinoma group was divided into two groups by the size of carcinoma (<1 cm and > or = 1 cm). The deltaCT value was 2.629 +/- 0.300 and 2.212 +/- 0.124 respectively (P<0.05) and the 2(-deltadeltaCT) value was 167.33 +/- 33. 823 and 221.69 +/- 18.843 respectively (P<0.01). The TSH level in carcinoma group and adenoma group had no statistical significance (P>0.05). CONCLUSION The expression of Runx2 mRNA was high in PTC, and was related to the size of carcinoma which was higher in bigger size carcinoma. The role of Runx2 may contribute to the formation of the micro-calcification and the development and progression in PTC and other malignant tumors, such as breast cancer, prostatic carcinoma and osteosarcoma.
Carcinoma ; metabolism ; pathology ; Carcinoma, Papillary ; Core Binding Factor Alpha 1 Subunit ; metabolism ; Female ; Humans ; Male ; Middle Aged ; RNA, Messenger ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Thyroid Cancer, Papillary ; Thyroid Neoplasms ; metabolism ; pathology

OBJECTIVE: To investigate the expression and clinical significance of Muc1, p63 protein in diffuse sclerosing variant of papillary thyroid carcinoma and conventional papillary thyroid carcinoma. METHOD: Immunohistochemistry (SP) was used to detect the expressions of Muc1, p63 protein in 30 samples of DSVPTC (experiment group) and 30 samples of CPTC (control group). Patients in two groups were matched in age, gender, tumor side, tumor size and date of diagnosis. RESULT: (1) The positive rate of Muc1 in DSVPTC and CPTC was 76.67% (23/30) and 53.33% (16/30) respectively, immunohistochemical staining expressed as brown or tan particles in the membrane or the cytoplasm,with a significant difference between the two groups (P < 0.05). The positive rate of p63 in DSVPTC and CPTC was 80% (24/30) and 43.33% (13/30) respectively, immunohistochemical staining expressed as a brown or tan particles in the muclei,with a significant difference between the two groups (P < 0.05). (2) Cervical lymph node metastasis rate in DSVPTC and CPTC was 50% (15/30) and 20% (6/30) respectively, with a significant difference between the two groups (P < 0.05). (3) In All cases,the positive rate of Muc1 in cervical lymph node metastasis group (21 cases) and without metastasis group (39 cases) was 85.71% (18/21) and 53.85% (21/39) respectively,with a significant difference between the two groups (P < 0.05); the positive rate of p63 was 95.24% (20/21) and 43.59% (17/39) respectively,with a significant difference between the two groups (P < 0.01). (4) Spearman rank correlation analysis showed that expression of Muc1 and p63 were positively correlated in both groups(r = 0.530,0. 386, P < 0.05). CONCLUSION (1) There are high expression of Muc1 and p63 protein in DSVPTC, and relatively low expression in CPTC, DSVPTC have a higher rate of cervical lymph node metastasis at the time of being diagnosed, compared to the CPTC. These results show that DSVPTC is a more biologically aggressive variant of the PTC. (2) Abnormal expression of Muc1 and p63 may be important to promote the progression and metastasis of PTC, thus they can be used as predictors of malignant behavior in PTC. (3) Muc1 and p63 may be synergistically promote proliferation and invasion metastasis of the PTC malignant cell.
Adolescent ; Adult ; Aged ; Carcinoma ; classification ; metabolism ; pathology ; Carcinoma, Papillary ; Female ; Humans ; Lymphatic Metastasis ; Male ; Membrane Proteins ; metabolism ; Middle Aged ; Mucin-1 ; metabolism ; Thyroid Cancer, Papillary ; Thyroid Neoplasms ; classification ; metabolism ; pathology ; Young Adult

OBJECTIVE: To investigate the expression of signal transducer and activator of transcription 3 (STAT3) and phosphorylated STAT3 (p-STAT3) protein in papillary thyroid carcinoma (PTC), and to explore the correlation and significance between the expression of STAT3, p-STAT3 and epithelial-mesenchymal transition (EMT). METHOD: The expression of STATS3. p-STAT3, E-cadherin and Vimentin protein in 56 cases of PTC specimens and adjacent normal tissues specimens ware detected by immunohistochemistry. The correlation of the expression of STATS, p-STAT3, E-cadherin and Vimentin protein in PTC with clinicopathological characteristics was analyzed. RESULT: The positive rates of STAT3, p-STAT3 in PTC tissue were significantly higher than those in adjacent normal tissues specimens respectively (P < 0.01). The positive rates of E-cadherin in PTC tissues were remarkably lower, compared to adjacent normal tissues specimens (P < 0.01), however the positive rates of Vimentin in PTC tissues were remarkably higher, compared to adjacent normal tissues specimens (P < 0.01). The expression of STAT3, p-STAT3, E-cadherin and Vimentin protein were associated with lymph node metastasis and clinical stage (all P < 0.05). The expression of STAT3 and p-STAT3 was negatively correlated with E-cadherin expression (r = -0.494 and r = -0.364, P < 0.01, respectively), but positively with Vimentin expression (r = 0.533 and r = 0.377, P < 0.01, respectively) in PTC tissues. CONCLUSION PTC tissues have STAT3 protein activation and EMT phenotype, as were all correlated significantly with PTC invasion and metastasis. STAT3 signaling pathway activation might mediate EMT and then promote PTC invasion and metastasis.
Adult ; Antigens, CD ; Cadherins ; metabolism ; Carcinoma ; metabolism ; pathology ; Carcinoma, Papillary ; Epithelial-Mesenchymal Transition ; Female ; Humans ; Male ; Middle Aged ; Phosphorylation ; STAT3 Transcription Factor ; metabolism ; Thyroid Cancer, Papillary ; Thyroid Neoplasms ; metabolism ; pathology ; Vimentin ; metabolism

OBJECTIVE: In order to assess the significance of Ezrin and E-Cadherin in invasion and metastasis of papillary thyroid carcinoma (PTC). METHOD: The 81 paraffin-embedded specimen PTC and normal thyroid tissues adjacent to cancer operated from 2005 to 2008 were tested. The expressions of Ezrin, E-Cadherin in the specimen were assayed by immunohistochemical staining. The correlation between the 2 factors (Ezrin, E-Cadherin) and clinical features of PTC such as gender, age, tumor size, invasion, lymph node metastasis, and TNM stage were studied. RESULTS: The rates of immuno-positive staining Ezrin in PTC was higher than the normal thyroid tissues adjacent to cancer with significant difference respectively (chi2 = 61.691, P < 0.01). The rates of immuno-positive staining E-Cadherin in PTC was lower than the normal thyroid tissues adjacent to cancer with significant difference respectively (chi2 = 64.241, P < 0.01). The expressions of Ezrin and E-cadherin were related to age, invasion, lymph node metastasis, and TNM. There was correlation between the expressions of Ezrin and E-Cadherin in PTC (r = -0.471, P < 0.01). CONCLUSION The high expressions of Ezrin and the low expression of E-Cadherin were related to invasion and metastasis in PTC. Between the 2 factors has cooperative. Combined detection of the expression of Ezrin and E-Cadherin in PTC might served as invasion, metastasis and important prognostic predictors.
Adult ; Aged ; Antigens, CD ; Cadherins ; metabolism ; Carcinoma ; metabolism ; pathology ; Carcinoma, Papillary ; Cytoskeletal Proteins ; metabolism ; Female ; Humans ; Male ; Middle Aged ; Neoplasm Invasiveness ; Neoplasm Metastasis ; Prognosis ; Thyroid Cancer, Papillary ; Thyroid Neoplasms ; metabolism ; pathology ; Young Adult