This study aimed to elucidate the mechanism by which SHP2 binding to PD-1 on T cells induces T cell exhaustion via the regulation of the AMPK/STAT3/EGR1/BRD4/NFAT2 signaling pathway, and its functional implications in thyroid cancer. First, single-cell RNA sequencing analysis was performed to characterize the tumor immune microenvironment, with a specific focus on the functional status of CD4 + regulatory T cells (Tregs). Bioinformatics analyses of TCGA and GTEx datasets were then conducted to evaluate the differential expression of SHP2, STAT3, BRD4, and NFAT2. Finally, the effects of SHP2 overexpression, PD-1 knockdown, and BRD4 knockdown on tumor growth and T cell function were examined using a nude mouse tumor xenograft model and in vitro cell culture assays. Multiple techniques, including Western blot, ELISA, flow cytometry, scratch wound assay, and Transwell assay, were applied to verify the regulation of the AMPK/STAT3/EGR1/BRD4/NFAT2 signaling pathway by SHP2-PD-1 binding and its impacts on T cell exhaustion and thyroid cancer cell phenotypes. Single-cell analysis demonstrated that CD4 + Tregs in papillary thyroid carcinoma displayed significantly elevated exhaustion and proliferation scores relative to normal thyroid tissues. In addition, pathway enrichment and pseudotime trajectory analyses revealed that Tregs within the tumor microenvironment were progressively transitioning toward a severe exhaustion state, driven by the upregulation of core signaling molecules. Bioinformatics analysis showed that SHP2, STAT3, BRD4, and NFAT2 were markedly overexpressed in thyroid cancer tissues, and high SHP2 expression was positively correlated with T cell exhaustion markers (e.g., TIM3, LAG3, TIGIT). In the nude mouse xenograft model, SHP2 overexpression significantly promoted tumor growth, while PD-1 or BRD4 knockdown inhibited tumor progression. In vitro cellular assays further confirmed that SHP2 interaction with PD-1 triggered T cell exhaustion through the AMPK/STAT3/EGR1/BRD4/NFAT2 signaling cascade, thereby modulating thyroid cancer cell migration, invasion, and apoptosis. SHP2–PD-1 interaction in T cells elicits T cell exhaustion via modulation of the AMPK/STAT3/EGR1/BRD4/NFAT2 signaling pathway, thereby accelerating thyroid cancer progression. These findings identify a novel potential therapeutic target for thyroid cancer immunotherapy.

Objective:To observe the expression level of circular RNA hsa_circ_0007813 in breast cancer tissues and its relationship with clinicopathologic features and survival rate of patients.Methods:A total of 132 cases of breast cancer tissue microarray were collected in Shanghai Outdo Biotech as the research objects. Fluorescence in situ hybridization (FISH) was used to detect the expression of hsa_circ_0007813, and to analyze the relationship between the expression of hsa_circ_0007813 and the clinicopathological characteristics and prognosis of patients. Chi-square test was used to analyze the relationship between hsa_circ_0007813 and clinicopathological features. Kaplan-Meier method was used to evaluate the overall survival rate of patients. Cox regression model was used for multivariate analysis.Results:FISH results showed that the positive rate of hsa_circ_0007813 expression in breast cancer tissues was 71.21% (94/132), which was significantly higher than that in adjacent tissues [28.79% (38/132), χ2= 47.515, P<0.05]. The positive rate of hsa_circ_0007813 expression in breast cancer tissues of patients with pathological stages Ⅲ-Ⅳ was 69.88% (58/83), which was significantly higher than 51.02% (25/49) in breast cancer tissues of patients with stage Ⅰ-Ⅱ ( χ2= 4.695, P<0.05). The positive rate of hsa_circ_0007813 expression in patients with lymph node metastasis was 81.01% (64/79), which was significantly higher than 56.60% (30/53) in patients without lymph node metastasis ( χ2= 9.219, P<0.05). Kaplan-Meier survival analysis showed that the survival rate of patients in high hsa_circ_0007813 expression group was 73.40% (69/94), which was significantly lower than that in low hsa_circ_0007813 expression group [97.37% (37/38), χ2=8.930, P<0.05]. Multivariate regression analysis showed clinical stage [hazard ratio ( HR)=3.246, 95% confidence interval ( CI): 1.273-8.368, P<0.05] and expression level of hsa_circ_0007813 ( HR=3.540, 95% CI: 1.053-11.903, P<0.05) were independent risk factors affecting the overall survival rate of breast cancer patients. Conclusion:Circular RNA hsa_circ_0007813 was highly expressed in breast cancer tissue, which was significantly related to the pathological stage, lymph node metastasis and survival rate of patients.

AbstractBackground Papillary thyroid cancer (PTC) is the most common endocrine malignant tumour. The purpose of this study was to explore the potential molecular mechanism of circRNA regulating immune-related mRNA through sponge miRNA in the occurrence and immune regulation of PTC.Methods All data were downloaded from public databases, such as GEO, Immport and TCGA. Differentially expressed (DE) mRNAs (DEmRNAs), DEmiRNAs and DEcircRNAs were identified using metaMA and limma packages. Subsequently, immune-related DEmRNAs were screened, and circRNA-miRNA-mRNA (ceRNA) regulatory network was constructed. In addition, functional annotation, protein-protein interaction (PPI) network construction, immune cell infiltration analysis and Pearson correlation analysis were performed. Finally, qRT-PCR validation and cell experiments were also performed.Results In total, 2962 DEmRNAs, 78 DEmiRNAs and 51 DEcircRNAs were obtained. Subsequently, 195 immune-related DEmRNAs were obtained based on Immport database. Cytokine-cytokine receptor interaction was the only signalling pathway obtained in KEGG analysis. Then, 8 hub immune-related DEmRNAs were identified based on PPI network and CytoHubba plug-in. Subsequently, ceRNA sub-network containing hub immune-related DEmRNAs was extracted from ceRNA regulatory network. In ceRNA sub-network, hsa_circ_0082182-hsa-miR-18b-5p-FGF1/PDGFC, hsa_circ_0016404-hsa-miR-1275-FGF1/CTSB/IL13RA1, hsa_circ_0070100-hsa-miR-27a-3p/hsa-miR-27b-3p-TGFBR3, hsa_circ_0060055/hsa_circ_0038718-hsa-miR-150-3p-CXCL14, hsa_circ_0030427/hsa_circ_0002917-hsa-miR-22-3p-BMP7 and hsa_circ_0030427/hsa_circ_0002917-hsa-miR-125a-5p-LIFR axes were identified. Moreover, FGF1, PDGFC, CTSB, IL13RA1, TGFBR3, CXCL14, BMP7, LIFR, hsa-miR-125a-5p, hsa-miR-1275, hsa-miR-150-3p, hsa-miR-18b-5p and hsa-miR-27b-3p were also found to have good diagnostic accuracy and may be potential novel diagnostic markers for PTC. XCell analysis showed that the levels of immune cell infiltration (including Tregs, HSC, DC and Monocytes) were significantly different between the PTC and the control groups. Knockdown of the expression of hsa_circ_0082182 significantly inhibits the activity, proliferation, migration and invasion of TPC-1 cells.Conclusion Several circRNA-miRNA-mRNA axes identified in this study may be related to the occurrence, progression and survival of PTC. This lays a theoretical foundation for further understanding the molecular mechanism of PTC, and also contributes to clinical management and research.

Objective:To observe the effect of long non-coding RNA small nucleolar RNA host gene 22 (SNHG22) targeting microRNA (miR)-27a-3p/insulin-like growth factor-1 (IGF-1) axis on cell invasion in triple negative breast cancer cells.Methods:A total of 30 cases of triple negative breast cancer and adjacent tissues collected in the Second Hospital of Hebei Medical University from June 2019 to March 2022 were selected as the research objects. Real-time polymerase chain reaction (PCR) method was used to observe the expression level of SNHG22 and miR-27a-3p in triple negative breast cancer and adjacent tissues. The SNHG22 overexpression plasmid and miR-27a-3p mimics were transfected into triple negative breast cancer MDA-MB-231 cells, respectively, and the effect of SNHG22 and miR-27a-3p on the invasion ability of triple negative breast cancer cells was observed by the cell invasion experiment. The relationship between SNHG22 and miR-27a-3p and the downstream target gene of miR-27a-3p was analyzed using bioinformatics software and double luciferase reporter gene. After co-transfection of miR-27a-3p mimics and miRNA negative control (miR-NC) with SNHG22, the effect of up-regulation of miR-27a-3p expression on the invasion ability of MDA-MB-231 cells overexpressing SNHG22 was observed. T-test was used to compare the two groups. Results:The results of real-time PCR experiment indicated that the expression level of SNHG22 in triple negative breast cancer tissues (3.184±1.800) was significantly higher than that in adjacent tissues (1.295±0.659, t=5.395, P<0.05), and the expression level of miR-27a-3p (0.547±0.414) was obviously lower than that of adjacent tissues (1.521±0.845, t=5.623, P<0.05). The number of invasive cells was significantly increased after transfection with SNHG22 overexpression plasmid (406.500±9.492) as compared with that in the control group (172.000±14.142, t=67.000, P<0.05), while the number of invasive cells in miR-27a-3p mimics transfection group (139.000±19.789) was significantly reduced as compared with that in the control group (391.000±21.213, t=252.000, P<0.05). Bioinformatics prediction and luciferase reporter gene analysis showed that miR-27a-3p could combine with SNHG22 to reduce the luciferase activity of cells (0.415±0.054 vs. 1.014±0.106, t=16.360, P<0.05), while miR-27a-3p could combine with the 3′untranslated region of IGF-1 to reduce the luciferase activity of cells (0.367±0.049 vs. 1.015±0.021, t=12.860, P<0.05). The results of rescue experiment indicated that the number of invasive cells in SNHG22+ miR-27a-3p mimics group (186.500±14.849) was significantly reduced as compared with that in SNHG22+ miR-NC group (345.000±24.061), and the difference was statistically significant ( t=24.380, P<0.05). Conclusion:Long chain non coding RNA SNHG22 can regulate the invasion ability of triple negative breast cancer cells by targeting miR-27a-3p/IGF-1 axis.

Objective:To observe the effect of long non-coding RNA (lncRNA) taurine upregulated gene 1 (TUG1) targeting microRNA (miR)-145/zinc-finger E-box binding homeobox 1 (ZEB1) on the invasion of papillary thyroid carcinoma (PTC) cells.Methods:Real-time polymerase chain reaction (PCR) method was used to detect the expression level of lncRN TUG1 and miR-145 in PTC and the adjacent tissues. The lncRNA TUG1 shRNA and miR-145 mimics were transfected into papillary thyroid carcinome cells-1(TPC-1) cells, respectively, and the effect of lncRNA TUG1 and miR-145 on the invasion ability of PTC cells was observed by the Transwell assay. The bioinformatics and dual luciferase reporter gene used to analyze the relationship between lncRNA TUG1 and miR-145 and the target gene of miR-145. Western blotting was used to verify the influence of lncRNA TUG1 and miR-145 on the expression level of target protein. Two-tailed Student′s t-test was used to evaluate the differences between two groups. Results:Real-time PCR showed that the expression level of lncRNA TUG1 in PTC tissues (4.43±0.07) was significantly higher than that in adjacent tissues (1.13±0.06, t=3.124, P<0.05), and the expression level of miR-145 (0.39±0.08) was lower than that in adjacent tissues (1.08±0.04, t=2.937, P<0.05); The number of invasive cells in TPC-1 cells transfected with lncRNA TUG1 shRNA (292.4±48.2) and miR-145 mimics (234.2±42.4) was lower than that in control group (452.9±50.8, 439.4±39.8, t=3.205, P<0.05; t=3.186, P<0.05); The results of bioinformatics and dual-luciferase reporter gene analysis showed that lncRNA TUG1 could bind to miR-145, and ZEB1 was the target gene of miR-145. Compared with the respective control cells (1.17±0.12, 1.05±0.12), the expression level of ZEB1 protein in lncRNA TUG1 shRNA group was down-regulated (0.33±0.12), and the difference was statistically significant ( t=3.320, P<0.05), and the protein expression level in miR-145 mimics transfected group was down-regulated (0.46±0.11), and the difference was statistically significant ( t= 3.450, P<0.05). Conclusion:lncRNA TUG1 can target miR-145/ZEB1 to regulate the invasion ability of PTC cells.

Objective:To observe the effect of sorafenib on ferroptosis in thyroid cancer TPC-1 cells, and to provide a new strategy for the treatment of thyroid cancer.Methods:Thyroid cancer TPC-1 cells were cultured in vitro, treated and divided into sorafenib group and negative control group, and cell counting kit-8 (CCK-8) assay was used to detect cell viability; Dichlorofluorescein (DCFH-DA) probe was used to detect reactive oxygen species (ROS) production; Western blotting experiment was used to detect the protein expression levels of ACSL4, SLC7A11 and GPX4; Malondialdehyde (MDA) kit was used to detect the content of MDA in thyroid cancer cells. Analysis of variance was used for the mean comparison between multiple groups, and t-test was used for the comparison between the two groups. Results:The cell absorbance value of sorafenib intervention group (1.29±0.17) was significantly lower than that of control group (1.86±0.23, t=2.695, P<0.05); The fluorescence intensity of ROS in TPC-1 cells treated with sorafenib (1.833±0.293) was significantly higher than that in the control group (0.825±0.186, t=2.801, P<0.05); Western blotting experiments showed that the expression level of ACSl4 protein (0.954±0.091) after sorafenib intervention was higher than that in the control group (0.442±0.132, t=5.314, P<0.05), and the protein expression level of SLC7A11 (0.293±0.087) was significantly lower than that in the control group (0.897±0.098, t=7.916, P<0.05), the protein expression level of GPx4 (0.325±0.051) was also lower than that of control cells (0.893±0.069, t=9.325, P<0.05). The results of MDA detection showed that the expression level of MDA in sorafenib intervention group (30.661±3.192) was significantly higher than that in control group (19.285±1.406, t=5.901, P<0.05). Conclusion:Sorafenib can promote the ferroptosis process of thyroid cancer cells and accelerate the death of thyroid cancer cells.

Objective:To observe the effect of microRNA (miRNA, miR)-630 in thyroid cancer (TC) cells on the expression of the epithelial-mesenchymal transition (EMT) marker Slug, and its influence on cell invasion.Methods:A total of 40 cases of papillary TC and their para-cancerous specimens were collected from the Department of General Surgery of Hebei Medical University from January 2018 to December 2019. The real-time quantitative polymerase chain reaction (Real-time PCR) was used to detect the expression level of miR-630 in TC and its adjacent tissues. After transfecting miR-630 mimics into human TC cell line TPC-1, the expression level of Slug gene was detected by Real-time PCR, and the regulatory effect of miR-630 on Slug gene was observed by double luciferase reporter gene assay. The overexpression Slug plasmid and miR-630 mimics were co-transfected into TPC-1 cells. Transwell cell invasion assay was used to observe the effect of miR-630 and Slug expression on the invasion of TC cells.Results:Real-time PCR results showed that the expression level of miR-630 in papillary TC tissues was (1.13±0.07), significantly lower than that in adjacent tissues (5.33±0.08) with the difference being statistically significant ( t=15.472, P<0.05), and miR-630 mimics up-regulated miR-630 in papillary TC cells, and Slug mRNA expression level (0.48±0.09) was also significantly lower than that in control group (1.03±0.07, t=7.132, P<0.05). The luciferase activity of miR-630 mimics and Slug 3′wild-type co-transfection group was significantly lower than that of control group (0.38±0.03, t=13.021, P<0.05). However, in the co-transfected group of miR-630 mimics and Slug 3′mutant, the luciferase activity did not change significantly (1.08±0.02 vs. 1.02±0.03, t=1.122, P>0.05). The number of invasive cells in mir-630 mimics group (241.3±42.4) was significantly less than that in control group (467.5±51.7, t=3.214, P<0.05). Compared with the negative control group (418.3±31.2), the invasion ability of the cells co-transfected with miR-630 mimics and Slug overexpression plasmids (411.6±28.3) was not statistically different ( t=1.131, P>0.05). Conclusion:miR-630 can inhibit the invasion ability of TC cells by targeting Slug.

Introduction Thyroid cancers are the most common malignancy of the endocrine system. Increasing evidence has suggested potential roles for cancer susceptibility candidate 15 (CASC15) in thyroid cancer. Papillary thyroid cancer (PTC) accounts for 80% of thyroid cancer, posing a great threat to public health. The present study aims to investigate the potential roles of CASC15 in PTC. Material and methods PTC tissues were obtained from patients diagnosed with PTC at the Second Hospital of Hebei Medical University from October 2016 to January 2018. qRT-PCR was performed to examine the mRNA levels of CASC15, Bcl-2/BAX, E-cadherin, caspase-3, and PI3K/AKT. Western blotting was applied to determine the protein levels of PI3K/AKT, CASC15, E-cadherin, BAX, Bcl-2 and caspase-3. CCK-8 assays were used to determine the viability of BCPAP and K1 cancer cells. Wound healing and transwell assays were conducted to evaluate the migration and invasion abilities of BCPAP and K1 cells. Results The results showed that CASC15 was upregulated in thyroid cancer tissues. Moreover, CASC15 in BCPAP and K1 was decreased by CASC15 siRNA. The progression, migration and invasion of BCPAP cells were significantly decreased after transfection with CASC15 siRNA, which was consistent with that in K1 cells. Furthermore, CASC15 siRNA treatment decreased the level of Bcl-2, and PI3K/AKT, and increased the expression of E-cadherin, BAX, and caspase-3. Conclusions Downregulated CASC15 inhibited the proliferation, migration and invasion of thyroid cancer. Therefore, CASC15 may be an oncogene in thyroid cancer and may serve as a target marker for the treatment of thyroid cancer.

The purpose of this study was to explore the potential role of HOTAIR in thyroid cancer carcinogenesis. We found that HOTAIR was unregulated in human thyroid cancer and inversely correlated with miR-1. Functional assays indicated HOTAIR regulates miR-1 directly in thyroid cancer cells. We also revealed that HOTAIR promotes the processes of thyroid cancer cell malignancy through regulation of microRNA-1 (miR-1). Furthermore, we showed that HOTAIR could regulate a downstream target of miR-1, CCND2, in a miR-1-mediated manner. In addition, we also proved, using a tumor formation assay in nude mice, that silencing HOTAIR inhibited tumor formation in vivo. Therefore, our study demonstrated that HOTAIR promotes the development and progression of thyroid cancer through inhibition of microRNA-1 and activation of CCND2.

目的 探讨甲状腺全切术后血钙、血镁、血磷和血清甲状旁腺激素(parathyroid hormone,PTH)水平的变化及意义.方法 选取甲状腺全切术患者129例(甲状腺乳头状癌82例,结节性甲状腺肿和甲状腺腺瘤47例),监测所有患者术前和术后30 min、1d、3d的血钙、血镁、血磷和血PTH水平;并分析患者性别、年龄、术前促甲状腺激素(thyroid stimulating hormone,TSH)水平、术中识别甲状旁腺数和病理类型与术后甲状旁腺损伤所致低钙血症间的关系.结果 不同甲状腺全切术组术后血钙、血镁和PTH水平均呈下降趋势,且血钙和血镁水平以术后1d下降最为明显,PTH水平在术后30 min即出现明显下降.血钙水平在不同甲状腺全切术组组间、不同时点间差异有统计学意义(P＜0.05),在组间·不同时点间交互作用差异无统计学意义(P＞0.05).血镁和PTH水平在不同甲状腺全切术组组间、不同时点间以及组间·不同时点间交互作用差异均有统计学意义(P＜0.05).血磷在不同甲状腺全切术组组间、不同时点间以及组间·不同时点间交互作用差异均无统计学意义(P＞0.05).129例甲状腺全切术患者中又分为血钙正常亚组77例(59.7％),低钙血症无症状亚组31例(24.0％),低钙血症有症状亚组21例(16.3％).低钙血症有症状亚组和低钙血症无症状亚组乳头状癌检出率高于血钙正常亚组(P＜0.05).血钙正常亚组、低钙血症有症状亚组和低钙血症无症状亚组性别、年龄、术前TSH水平以及术中识别甲状旁腺数差异无统计学意义(P＞0.05).结论 手术范围和甲状腺癌可能是术后甲状旁腺损伤的影响因素.甲状腺术后血PTH监测较血钙更敏感.甲状腺术后低钙血症多合并低镁血症,补钙同时应补镁.

Objective To study the expression of Gli1 protein and vascular endothelial growth factor (VEGF)-C in papillary thyroid carcinoma and their relationship with cervical lymph node metastasis.Methods The expression of Gli1 protein and VEGF-C were investigated by immunohistochemical EnVision method in 67 cases of papillary thyroid carcinoma and 32 cases of nodular goiter pathological specimen tissues.Nascent microvascular and micro-lymphatic of all pathological speimen tissues were examined by immunohistochemical monoclonal antibody CD34 and D2-40 staining assay respectively,and the microvascular density (MVD) and micro-lymphatic vessel density (MLVD) were calculated.The relationship between the positive expression of Gli1 protein and MVD,between the positive expression of VEGF-C and MLVD,and their relationship with cervical lymph node metastasis were analyzed.Results The positive rates of Gli1 protein,VEGF-C and MVD,MLVD were significantly higher in papillary thyroid carcinoma than those in nodular goiter [67.16％(45/67) vs.43.75％(14/32),70.15％(47/67) vs.31.25％(10/32),23.14 ± 2.06 vs.2.14 ± 0.31,13.36 ± 1.32 vs.3.53 ± 0.65,P ＜ 0.05].The positive expression of Gli1 protein was related to the patient' s age and the tumor size (P ＜ 0.05),while the positive expression of VEGF-C was not related to the patient' s age and the tumor size (P ＞ 0.05).The positive expression of Gli1 protein and VEGFC were significantly higher in TNM stage Ⅲ-Ⅳ of papillary thyroid carcinoma than those in TNM stage Ⅰ-Ⅱ (P ＜ 0.01) and also was significantly higher with cervical lymph node metastasis than without cervical lymph node metastasis (P ＜ 0.01).MVD and MLVD were significantly higher with cervical lymph node metastasis than without cervical lymph node metastasis (P ＜ 0.05).The positive expression of Gli1 protein was positively correlated with MVD (r =0.784,P＜ 0.01),the positive expression of VEGF-C was positively correlated with MLVD (r =0.529,P ＜ 0.01),the positive expression of Gli 1 protein was positively correlated with the positive expression of VEGF-C (r =0.586,P ＜0.01).Conclusions Gli1 protein which may participate in the formation of nascent microvascular is abnormally activated by the Hedgehog signaling pathway to express,VEGF-C which may be participate in the formation of nascent micro-lymphatic is mediated and started by the MAPK signaling pathways to express.Expressions of Gli1 protein is positively correlated with expressions of VEGF-C,therefore the Hedgehog signaling pathway may be associated with the MAPK signal pathway.Suppressing nascent microvascular and micro-lymphatic may become new target to blockingup papillary thyroid carcinoma cervical lymph node metastasis.