Papillary Thyroid Carcinoma Cell Proliferation Research Articles

ACAP3 negatively regulated by HDAC2 inhibits the malignant development of papillary thyroid carcinoma cells

ArfGAP with coiled-coil, ankyrin repeat and PH domains 3 (ACAP3) level has been confirmed to be downregulated in papillary thyroid carcinoma (PTC). Histone deacetylase inhibitors (HDACIs) have therapeutic effects on PTC. Accordingly, this study probed into the potential relation of HDACIs and ACAP3 in PTC. Expressions of ACAP3 and histone deacetylase 2 (HDAC2) in PTC were investigated by quantitative real-time polymerase chain reaction (qRT-PCR). The relationship between HDAC2 and ACAP3 was predicted by Pearson analysis. Cell functional assays (cell counting kit-8, transwell, wound healing and flow cytometry assays) and rescue assay were carried out to determine the effects of HDAC2/ACAP3 axis on biological behaviors of PTC cells. Expressions of apoptosis-, epithelial-mesenchymal transition-, Protein Kinase B (AKT)-, and P53-related proteins were measured by Western blot. ACAP3 level was downregulated in PTC tissues and cells. ACAP3 overexpression (oe-ACAP3) suppressed viability, proliferation, migration and invasion of PTC cells, facilitated apoptosis, downregulated the expressions of Protein Kinase B (Bcl-2) and N-cadherin, upregulated the expressions of Bcl-2 associated protein X (Bax) and E-cadherin, diminished the p-AKT/AKT ratio and elevated the p-p53/p53 ratio; however, ACAP3 silencing or HDAC2 overexpression (oe-HDAC2) did the opposite. HDAC2 negatively correlated with ACAP3. The tumor-suppressing effect of oe-ACAP3 in PTC was reversed by oe-HDAC2. Collectively, ACAP3 negatively regulated by HDAC2 suppresses the proliferation and metastasis while facilitating apoptosis of PTC cells.

MAZ promotes thyroid cancer progression by driving transcriptional reprogram and enhancing ERK1/2 activation

Papillary thyroid carcinoma (PTC) is the most common type of thyroid malignancies worldwide. Oncogenic transcription factors (TFs) drive transcriptional reprogramming and tumorigenesis. The myc-associated zinc finger protein (MAZ) is one of the Myc family TFs. The role of MAZ in PTC pathogenesis is still largely unknown. Here, we report that MAZ profoundly promotes proliferation of PTC cells ex vivo and in vivo through activating MAPK signaling. We firstly profiled gene expression of PTC cells after silencing of MAZ. BRAF, KRAS and LOC547 were identified as important target genes of TF MAZ. In particular, TF MAZ bound to the promoters of BRAF or KRAS and significantly increased their transcription and expression levels. Meanwhile, MAZ could noticeably elevate LOC547 transcription and expression as a TF. High levels of LOC547 relocated ACTN4 protein from the nucleus to the cytosol, improved protein-protein interactions between ACTN4 and EGFR in the cytosol, enhanced ERK1/2 phosphorylation, activated the MAPK signaling and, thus, promoted PTC progression. Our data identify a previously underappreciated MAZ-controlled transcriptional reprogram and ERK1/2 activation via BRAF, KRAS and LOC547. Our data illustrate that activation of the MAZ-controlled axis promotes thyroid tumorigenesis. These insights would advance our knowledge of the role of TFs in cancer development and highlight the potential of TFs as future targets for treatments against cancers.

Circ-METTL15 stimulates the aggressive behaviors of papillary thyroid cancer cells by coordinating the miR-200c-3p/XIAP axis.

Papillary thyroid carcinoma (PTC) is the most common form of thyroid cancer. The critical importance of circular RNA (circRNA) in a range of cancer types has been lately recognized. However, research on the functions of circRNAs in PTC has been limited thus far. Therefore, this research aimed at exploring the function and mechanism of circ-methyltransferase-like 15 (METTL15) in PTC cells. Quantitative measurements of circ-METTL15, miR-200c-3p, and X-linked inhibitor of apoptosis protein (XIAP) in PTC cells were conducted using reverse transcription-quantitative polymerase chain reaction or Western blot analysis. To investigate cell growth, cell counting kit-8 and colony formation tests were employed, apoptosis was analyzed using flow cytometry, and migration and invasion were studied through Transwell assays. The targeted binding sites between miR-200c-3p and circ-METTL15 or XIAP were predicted by starBase and then verified by dual luciferase reporter assay. circ-METTL15 and XIAP were upregulated in the PTC cells, while miR-200c-3p was downregulated. Downregulating circ-METTL15 or upregulating miR-200c-3p resulted in inhibited proliferation, migration, and invasion of PTC cells, while promoting apoptosis. miR-200c-3p was the downstream molecule of circ-METTL15, and XIAP was the direct target of miR-200c-3p. Forcing XIAP expression obstructed circ-METTL15 silencing to inhibit PTC cell activity. By coopting miR-200c-3p/XIAP, Circ-METTL15 stimulates aggressive behavior in PTC cells.

CircPHGDH downregulation decreases papillary thyroid cancer progression through miR-122-5p/PKM2 axis

BackgroundAlthough papillary thyroid carcinoma (PTC) has a favorable prognosis, it could affect patient life quality and become a serious threat because of invasion and metastasis. Many investigations have suggested that circular RNAs (circRNAs) are involved in different cancer regulations. Nevertheless, circRNAs role in invasive PTC remains unclear.MethodsIn the present investigation, next-generation sequencing was applied to explore abnormal circRNA expression. The expression of circRNA phosphoglycerate dehydrogenase (circPHGDH) in PTC cell lines and tissues were examined. Then, we investigated regulatory mechanism and circPHGDH downstream targets using bioinformatics analysis and luciferase reporting analysis. Then transwell migration, Cell Counting Kit-8 (CCK8) and 5-ethynyl-2’-deoxyuridine (EdU) assays were used for cells migration and proliferation analysis. In vivo metastasis and tumorigenesis assays were also employed to evaluate the circPHGDH role in PTC.ResultsThe data showcased that circPHGDH expression increased in both PTC cell lines and tissues, which suggested that circPHGDH functions in PTC progression. circPHGDH downregulation suppressed PTC invasion and proliferation in both in vivo and in vitro experiments. Bioinformatics and luciferase reporter results confirmed that both microRNA (miR)-122-5p and pyruvate kinase M2 subtype (PKM2) were downstream targets of circPHGDH. PKM2 overexpression or miR-122-5p suppression reversed PTC cell invasion and proliferation post silencing circPHGDH by restoring aerobic glycolysis.ConclusionTaken together, our research found that circPHGDH downregulation reduced PTC progression via miR-122-5p/PKM2 axis regulation mediated by aerobic glycolysis.

Tumor‑suppressive effects of Smad‑ubiquitination regulator 2 in papillary thyroid carcinoma.

Smad-ubiquitination regulator 2 (SMURF2) functions as a homolog of E6AP carboxyl terminus-type E3 ubiquitin ligase to regulate cell cycle progression and tumor growth factor expression. SMURF2 has been revealed to function as a tumor suppressor in a number of cancers; however, its function in papillary thyroid carcinoma (PTC) remains largely unknown. Therefore, the aim of the present study was to investigate the function of SMURF2 in PTC. Reverse transcription-quantitative PCR and western blotting were used to detect cellular expression of SMURF2 in vitro. After increasing or inhibiting the expression of SMURF2, MTT was used to detect the effect on tumor cell proliferation and Transwell assays were used to detect the effect on tumor cell migration and invasion. Finally, ELISA was used to detect the effects on glucose and glutamine metabolism in tumor cells and the findings revealed that SMURF2 was downregulated in PTC tissues. Moreover, SMURF2 inhibited the proliferation, invasion and migration of PTC cells, and promoted their apoptosis. Finally, SMURF2 inhibited cell glycolysis and glutaminolysis and affected metabolism in the PTC cell line, TPC-1. Thus, the findings of the present study suggest that SMURF2 may be a potential target in the treatment of PTC.

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.

MiR-374a/b-5p Suppresses Cell Growth in Papillary Thyroid Carcinoma Through Blocking Exosomal ANXA1-Induced Macrophage M2 Polarization.

Papillary thyroid carcinoma (PTC), comprising 85% of all thyroid cancers, is an epithelial malignancy. The potential for malignant transformation in normal cells by thyroid cancer cells via exosomal Annexin A1 (ANXA1) delivery is investigated in this study. Our aim is to determine the impact of PTC cells on macrophage polarization through exosomal ANXA1 secretion and its implications for tumor progression.Exosomes in PTC cells were examined using transmission electron microscopy, exosome labeling, and nanoparticle tracking analysis. Real-time quantitative polymerase chain reaction was employed to quantify gene expression levels. Protein levels were determined through Western blot analysis. The interplay between genes was assessed using luciferase reporter and RNA pull-down assays. Functional experiments were conducted to investigate PTC cell proliferation and apoptosis.Our findings reveal that ANXA1 promotes PTC cell proliferation and inhibits apoptosis. Exosomes derived from PTC cells were found to promote macrophage M2 polarization. ANXA1 stimulates M2 polarization through the activation of the PI3K/AKT pathway. MicroRNA-374a-5p (miR-374a-5p) and microRNA-374b-5p (miR-374b-5p) were identified as inhibitors of ANXA1 expression and PI3K/AKT pathway activity, thereby inhibiting macrophage M2 polarization. Furthermore, miR-374a-5p and miR-374b-5p were observed to suppress PTC cell proliferation through their regulatory action on ANXA1.Our study suggests that miR-374a/b-5p inhibits PTC cell growth by blocking the macrophage M2 polarization induced by exosomal ANXA1.

CircNRCAM up-regulates NRCAM to promote papillary thyroid carcinoma progression.

Papillary Thyroid Carcinoma (PTC) is the most prevalent subtype of Thyroid Carcinoma (THCA), a type of malignancy in the endocrine system. According to prior studies, Neural Cell Adhesion Molecule (NRCAM) has been found to be up-regulated in PTC and stimulates the proliferation and migration of PTC cells. However, the specific mechanism of NRCAM in PTC cells is not yet fully understood. Consequently, this study aimed to investigate the underlying mechanism of NRCAM in PTC cells, the findings of which could provide new insights for the development of potential treatment targets for PTC. Bioinformatics tools were utilized and a series of experiments were conducted, including Western blot, colony formation, and dual-luciferase reporter assays. The data collected indicated that NRCAM was overexpressed in THCA tissues and PTC cells. Circular RNA NRCAM (circNRCAM) was found to be highly expressed in PTC cells and to positively regulate NRCAM expression. Through loss-of-function assays, both circNRCAM and NRCAM were shown to promote the proliferation, invasion, and migration of PTC cells. Mechanistically, this study confirmed that precursor microRNA-506 (pre-miR-506) could bind with m6A demethylase AlkB Homolog 5 (ALKBH5), leading to its m6A demethylation. It was also discovered that circNRCAM could competitively bind to ALKBH5, which restrained miR-506-3p expression and promoted NRCAM expression. In summary, circNRCAM could up-regulate NRCAM by down-regulating miR-506-3p, thereby enhancing the biological behaviors of PTC cells.

PAFAH1B3 Regulates Papillary Thyroid Carcinoma Cell Proliferation and Metastasis by Affecting the EMT.

Thyroid carcinoma (TC) is currently the prevalent type of endocrine malignancy worldwide, having an incidence of around 15.5 per 100,000 people. However, the underlying mechanisms of TC tumorigenesis remain to be further elucidated. Performing the database analyses, Platelet-activating factor acetylhydrolase 1B3 (PAFAH1B3) was found to be dysregulated in several carcinomas and might trigger tumor occurrence as well as the progression of TC. Clinicopathological information of patients from our local validated cohort and The Cancer Genome Atlas (TCGA) cohort also confirmed this hypothesis. Our present research showed that elevated expression of PAFAH1B3 has a close association with worse behavior in papillary thyroid carcinoma (PTC). We utilized the small interfering RNA to obtain the PAFAH1B3-transfected PTC cell lines, including BCPAP, FTC-133, and TPC-1, and then further examined their biological function in vitro. Furthermore, gene set enrichment analysis suggested that PAFAH1B3 is implicated with epithelial-mesenchymal transition (EMT). Afterward, the western blotting assays aimed at EMT-related proteins were performed. In short, our results revealed that silencing PAFAH1B3 could hinder the capabilities of proliferation, migration, and invasion of PTC cells. Increasing expression of PAFAH1B3 might be of quintessence with lymph node metastasis by triggering EMT in PTC patients.

Circ-LDLRAD3/miR-655-3p/MAPK1 axis enhances cell migration and invasion in papillary thyroid carcinoma.

Papillary thyroid carcinoma (PTC) is a prevalent histological subtype of thyroid cancer, whose occurrence and development may be related to circRNA dysregulation. This research proposed to unravel circ-LDLRAD3-related mechanisms in PTC. First, circ-LDLRAD3, miR-655-3p .and MAPK1 levels in PTC were quantitatively measured. Then, plasmid vectors or oligonucleotides that interfere with circ-LDLRAD3, miR-655-3p, or MAPK1 were transfected into PTC cells, followed by the analysis of proliferation, apoptosis, migration, and invasion. Finally, the targeted binding sites between miR-655-3p and circ-LDLRAD3 or MAPK1 were predicted by starBase and experimentally verified. Statistically, PTC samples expressed high circ-LDLRAD3 and MAPK1 and low miR-655-3p. Knocking down circ-LDLRAD3 or enhancing miR-655-3p hindered PTC cell proliferation, migration, and invasion, and forced apoptosis. circ-LDLRAD3 bound to miR-655-3p to affect MAPK1 expression. Elevating MAPK1 rescued circ-LDLRAD3 knockdown-allowed obstruction of PTC cell growth. In conclusion, circ-LDLRAD3 stimulates PTC development by releasing miR-655-3p-targeted MAPK1.

P53-associated miRNAs repress lncRNA ZFAS1 to retard the proliferation of papillary thyroid carcinoma.

Thyroid carcinoma is the most frequent malignancy among endocrine-related tumours. Papillary thyroid carcinoma (PTC) is the main type of thyroid carcinoma, and almost 80% cases of thyroid carcinoma are diagnosed as PTC. The molecular mechanism underlying PTC progression is unclear. This study aims to investigate the potential mechanisms of zinc finger antisense 1 (ZFAS1) function in PTC. The expression of ZFAS1 and p53 was determined by quantitative polymerase chain analysis (qPCR) in PTC tissues derived from 20 PTC patients. Quantitative chromatin immunoprecipitation assay (qChIP) analysis was performed to validate the target of ZFAS1/p53 and miRNAs/p53. The Gene Expression Omnibus (GEO) dataset GSE94908 was analysed to obtain the differentially expressed p53-associated microRNAs (miRNAs). Luciferase assay validated the target of ZFAS1/miRNAs, and 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to determine the cell proliferation. The expression of ZFAS1 was up-regulated in the tissues derived from PTC patients, and the expression of ZFAS1 was negatively associated with p53 expression in PTC. The expression of ZFAS1 was significantly higher in the MDA-T120 cells harbouring mutant p53. We validated that ZFAS1 is a direct target of p53. In PTC cells, p53 directly repressed the ZFAS1 expression. In addition, we determined that miR-135b-5p and miR-193a-3p are directly induced by p53 in PTC cells. Interestingly, p53-targeted miR-135b-5p, miR-193a-3p, and miR-34b repressed the expression of ZFAS1 via the seed-matching sequences in the 3'-untranslated region (3'-UTR) of ZFAS1, and thereby suppressed PTC cell proliferation induced by ZFAS1. The oncogenic lncRNA ZFAS1 is directly repressed by p53 in PTC. p53-mediated miRNAs including miR-135b 5p, miR-193a-3p, and miR-34b repress ZFAS1 expression, and thereby inhibit the proliferation of PTC.

PIWIL1 Promotes Malignant Progression of Papillary Thyroid Carcinoma by Inducing EVA1A Expression.

Papillary thyroid carcinoma (PTC) is the most common subtype of thyroid cancer. Previous studies have reported on the ectopic expression of P-element-induced wimpy testis ligand 1 (PIWIL1) in various human cancers, but its role in PTC progression has not been investigated. In this study, we measured the expression levels of PIWIL1 and Eva-1 homolog A (EVA1A) in PTC using qPCR and WB. We performed a viability assay to evaluate PTC cell proliferation and used flow cytometry to investigate apoptosis. Moreover, we conducted a Transwell invasion assay to quantify cell invasion and assessed PTC growth in vivo using xenograft tumor models. Our findings showed PIWIL1 to be highly expressed in PTC and promote cell proliferation, cell cycle activity, and cell invasion, while suppressing apoptosis. Additionally, PIWIL1 accelerated tumor growth in PTC xenografts by modulating the EVA1A expression. Our study suggests that PIWIL1 contributes to the progression of PTC through EVA1A signaling, indicating its potential role as a therapeutic target for PTC. These results provide valuable insights into PIWIL1 function and may lead to more effective treatments for PTC.

FUS-Mediated CircFGFR1 Accelerates the Development of Papillary Thyroid Carcinoma by Stabilizing FGFR1 Protein.

Papillary thyroid carcinoma (PTC) is the most prevalent type of thyroid cancer and its incidence is rising globally. The molecular mechanisms of PTC progression remain unclear, hindering the development of effective treatments. This study focuses on hsa_circ_0008016 (circFGFR1), a circular RNA significantly up-regulated in PTC cells. Silencing circFGFR1 inhibited PTC cell proliferation and increased cell apoptosis, suggesting its role in PTC progression. The RNA-binding protein FUS was identified as a promoter of circFGFR1 formation. While circFGFR1 does not influence FGFR1 mRNA translation, it inhibits ubiquitination and degradation of FGFR1 protein, prolonging its half-life. CircFGFR1 also interacts with protein CBL, inhibiting CBL-mediated ubiquitination of FGFR1 proteins. Rescue assays confirmed circFGFR1 promotes PTC cell growth through mediating FGFR1. This study highlights the potential of circFGFR1 as a therapeutic target, offering insights into PTC's molecular mechanisms, and paving the way for novel treatment strategies.

Long non-coding RNA ACTA2-AS1 suppresses metastasis of papillary thyroid cancer via regulation of miR-4428/KLF9 axis

BackgroundMetastasis is the primary cause of recurrence and death in patients with papillary thyroid carcinoma (PTC). LncRNA ACTA2-AS1, a long non-coding RNA, acts as a tumor suppressor in multiple types of human malignancies, while the role of ACTA2-AS1 in PTC metastasis remains unclear.MethodsThe ACTA2-AS1 expression in PTC tissues was analyzed. The sponged roles of ACTA2-AS1 via miR-4428/KLF9 axis were identified using starBase tool. The function of ACTA2-AS1 in PTC was performed with in vitro and in vivo experiments. The correlation between DNA methylation and mRNA expressions of these gene in the TCGA dataset was explored.ResultsACTA2-AS1 expression was downregulated in PTC tissues without metastasis and further decreased in PTC tissues with lymph node metastasis compared with that in normal tissues. Functionally, the overexpression of ACTA2-AS1 inhibited the growth, proliferation, and invasion of PTC cells, whereas its depletion exerted opposite effect. In vivo, ACTA2-AS1 expression inhibited PTC metastasis. Furthermore, ACTA2-AS1 acted as a competing endogenous RNA for miR-4428, thereby positively regulating the expression of miR-4428 target gene, KLF9. Finally, miR-4428 overexpression enhanced invasive potential of PTC cells and significantly weakened the effects of ACTA2-AS1 on promotion and inhibition of KLF9 expression as well as invasive ability of PTC cells, respectively. In the TCGA dataset, the methylation level of ACTA2-AS1 was significantly correlated with its mRNA expression (r = 0.21, p = 2.1 × e−6).ConclusionsOur findings demonstrate that ACTA2-AS1 functions as a tumor suppressor in PTC progression at least partly by regulating the miR-4428-dependent expression of KLF9.Graphical abstract

MicroRNA-3653-3p inhibited papillary thyroid carcinoma progression by regulating CRIPTO-1.

Papillary thyroid carcinoma (PTC) is the most common endocrine malignant tumor and the metastasis of PTC often leads to unfavorable prognosis. Thus, the purpose of the current research was to mainly explore the role of miR-3653-3p in PTC progression. The expression level of miR-3653-3p in PTC was determined by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR), and Cell Counting Kit-8 (CCK-8) assay and colony formation assay were recruited to assess the ability of miR-3653-3p on cell proliferation. Next, transwell assay and Matrigel assay were involved to examine the ability of miR-3653-3p on cell migration and invasion. At last, Dual-Luciferase reporter assay and Western blotting were recruited to validate the down-stream target of miR-3653-3p. Results showed that miR-3653-3p was down-expressed in PTC, and upregulated miR-3653-3p inhibited cell proliferation, cell migration, and cell invasion in vitro. In addition, CRIPTO-1 was a downstream target of miR-3653-3p, and miR-3653-3p inhibited PTC progression via regulating CRIPTO-1. In sum, this research verifies that miR-3653-3p suppresses cell proliferation, migration, and invasion in PTC via regulating CRIPTO-1. These findings provide new insight into the underlying mechanism of PTC progression and may be useful in finding biomarkers and therapeutic targets of PTC.

LncRNA PFAR facilitates the proliferation and migration of papillary thyroid carcinoma by competitively binding to miR-15a

Papillary thyroid carcinoma (PTC) is type of aggressive tumor, with a markedly declined survival rate when distant metastasis occurs. It is of great significance to develop potential biomarkers to evaluate the progression of PTC. LncRNAs are recently widely claimed with biomarker value in malignant tumors. Herein, the role of LncRNA PFAR in PTC was investigated to explore potential prognostic marker for PTC. Compared to NTHY-ORI 3-1 cells, LncRNA PFAR was found markedly upregulated in PTC cell lines. In LncRNA PFAR knockdown TPC-1 cells, markedly declined cell viability, increased apoptotic rate, enhancive number of migrated cells, and elevated migration distance were observed, accompanied by a suppressed activity of the RET/AKT/mTOR signaling. In LncRNA PFAR overexpressed BCPAP cells, signally increased cell viability, declined apoptotic rate, reduced number of migrated cells, decreased migration distance, and increased tumor volume and tumor weight in nude mice xenograft model were observed, accompanied by an activation of the RET/AKT/mTOR signaling. The binding site between LncRNA PFAR and miR-15a, as well as miR-15a and RET, was confirmed by the dual luciferase reporter assay. The FISH study revealed that LncRNA PFAR was mainly located in the cytoplasm. Furthermore, the impact of the siRNA targeting LncRNA PFAR against the growth and migration of PTC cells was abolished by the inhibitor of miR-15a or SC79, an activator of AKT/mTOR signaling. Collectively, LncRNA PFAR facilitated the proliferation and migration of PTC cells by mediating the miR-15a/RET axis.

Progranulin promoted the proliferation, metastasis, and suppressed apoptosis via JAK2-STAT3/4 signaling pathway in papillary thyroid carcinoma

BackgroundProgranulin (PGRN), a glycoprotein secreted by a wide range of epithelial cells and plays an important role in inflammatory mechanisms and tumor progression. In this study, the expression, and functions of PGRN in papillary thyroid carcinoma (PTC) was examined to explore the potential pathogenesis of PTC.MethodsWestern blotting and qRT-PCR were used to detect the relationship between PGRN expression and clinicopathological characteristics of patients with PTC. PTC cell lines with PGRN overexpression and with PGRN knockdown were established to explore their effects on the biological behavior. Western blotting was used to detect the changes of relevant molecules and JAK2-STAT3/4 signaling pathway. Moreover, rescue experiments validated the involvement of the JAK2-STAT3/4 signaling pathway. And statistical analyses were analyzed using SPASS 21.0 and graph generation were performed using GraphPad Prism 8.0.ResultsPGRN was overexpressed in PTC tissue and increased by 75% at mRNA level and 161% at relative protein level in the patients with lymph node metastasis compared to without lymph node metastasis. Besides, PGRN regulated and promoted PTC cell proliferation, migration, invasion, and inhibited cell apoptosis. With PGRN overexpressed, relevant molecules including the expression of BCL2/BAX, BCL2/BAD, CyclinD1, MMP2, vimentin and N-cadherin were increased, the expression level of E-cadherin was decreased, and the phosphorylation of JAK2 and STAT3/4 were increased. JAK inhibitor (JSI-124) rescued these changes of PTC cells induced by overexpressed PGRN.ConclusionsThese findings revealed that PGRN promote the progression of PTC through the JAK2-STAT3/4 pathway, and PGRN could be served as a potential therapeutic target for PTC.

CRISPR/Cas9-mediated high-mobility group A2 knockout inhibits cell proliferation and invasion in papillary thyroid carcinoma cells

PurposeMetastasis and recurrence are the prognostic risk factor in patients with thyroid carcinoma. High-mobility group A2 (HMGA2) protein plays a crucial role in papillary thyroid carcinoma (PTC) metastasis. The aim of this study was to investigate the mechanisms underlying the HMGA2 effect on PTC cell proliferation and invasion. Materials and methodsWe used the CRISPR/Cas9 system to perform knockout of the HMGA2 gene in the human PTC cell line TPC-1. The knockout monoclonal cells were screened and verified by PCR analysis and genomic sequencing. Cell proliferation was examined after the knockout of the HMGA2 gene using cell counting kit-8 (CCK-8) assays. Furthermore, cell migration and invasion after the knockout were examined by cell scratch tests. Additionally, the changes in cell cycle and apoptosis after the knockout were detected by flow cytometry. ResultsThe results of the PCR analysis and the genomic sequencing confirmed that the human PTC TPC-1 ​cell line with knockout of HMGA2 gene was successfully established. The knockout of the HMGA2 gene significantly reduced the cell proliferation, growth, and invasion. Meanwhile, the knockout of the HMGA2 gene delayed the conversion of the G2/M phase and promoted cell necrosis. ConclusionThe CRISPR/Cas9-mediated HMGA2 knockout in the TPC-1 ​cell line inhibited cell proliferation and invasion, which might be due to the blockage of the cell cycle in the G2/M phase and the promotion of cell necrosis.

Regulatory role of Chitinase 3-like 1 gene in papillary thyroid carcinoma proved by integration analyses of single-cell sequencing with cohort and experimental validations

Papillary thyroid carcinoma (PTC) is one of the most common thyroid carcinomas. The gross extrathyroidal extension and extensive metastases of PTC lead to high rates of recurrence and poor clinical outcomes. However, the mechanisms underlying PTC development are poorly understood. In this study, using single-cell RNA sequencing, the transcriptome profiles of two PTC patients were addressed, including PTC1 with low malignancy and good prognosis and PTC2 with high malignancy and poor prognosis. We found that epithelial subcluster Epi02 was the most associated with the malignant development of PTC cells, with which the fold change of Chitinase 3-like 1 (CHI3L1) is on the top of the differentially expressed genes between PTC1 and PTC2 (P < 0.001). However CHI3L1 is rarely investigated in PTC as far. We then studied its role in PTC with a series of experiments. Firstly, qRT-PCR analysis of 14 PTC patients showed that the expression of CHI3L1 was positively correlated with malignancy. In addition, overexpression or silencing of CHI3L1 in TPC-1 cells, a PTC cell line, cultured in vitro showed that the proliferation, invasion, and metastasis of the cells were promoted or alleviated by CHI3L1. Further, immunohistochemistry analysis of 110 PTC cases revealed a significant relationship between CHI3L1 protein expression and PTC progression, especially the T (P < 0.001), N (P < 0.001), M stages (P = 0.007) and gross ETE (P < 0.001). Together, our results prove that CHI3L1 is a positive regulator of malignant development of PTC, and it promotes proliferation, invasion, and metastasis of PTC cells. Our study improves understanding of the molecular mechanisms underlying the progression of PTC and provides new insights for the clinical diagnosis and treatment of PTC.

LINC00641 impeded the malignant biological behaviors of papillary thyroid carcinoma cells via interacting with IGF2BP1 to reduce GLI1 mRNA stability.

Dysregulation of long intergenic non-protein coding RNA 00,641 (LINC00641) is associated with the malignancy progression of multiple cancers, including thyroid carcinoma. The current study aimed to determine the role of LINC00641 in papillary thyroid carcinoma (PTC) and the underlying mechanism. We found that LINC00641 was downregulated in PTC tissues and cells(p < 0.05), and overexpression of LINC00641 inhibited PTC cell proliferation and invasion, and induced apoptosis(p < 0.05), while silencing LINC00641 promoted the proliferation and invasion in PTC cells, and inhibited cell apoptosis(p < 0.05). Furthermore, we found that Glioma-associated oncogene homolog 1 (GLI1) expression was negatively correlated with LINC00641 expression in PTC tissues (r2 = 0.7649, p < 0.0001), and silencing GLI1 inhibited PTC cell proliferation and invasion, and induced apoptosis(p < 0.05). Meanwhile, RNA immunoprecipitation (RIP) and RNA pull-down assays confirmed that insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1) bound to LINC00641 as an RNA binding protein, and overexpression of LINC00641 destabilized GLI1 mRNA by competitively binding to IGF2BP1. Rescue experiments revealed that overexpression of GLI1 restored the inhibitory effect of LINC00641 overexpression on activation of the AKT pathway, as well as PTC cell proliferation and invasion, and counteracted the induction of cell apoptosis by LINC00641 overexpression. Finally, in vivo experimental results showed that overexpression of LINC00641 markedly suppressed tumor growth and reduced expression of GLI1 and p-AKT in xenograft tumor mice(p < 0.05). In summary, this study highlighted that LINC00641 plays a critical role in the malignant biological progression of PTC by regulating the LINC00641/IGF2BP1/GLI1/AKT signaling pathway, which may serve as a potential therapeutic target for PTC.