Biomarker Discovery for Early Diagnosis of Papillary Thyroid Carcinoma Using High-Throughput Enhanced Quantitative Plasma Proteomics.

The incidence of thyroid cancer (TC) has been increasing over the last 50 years worldwide. A higher rate of overdiagnosis in indolent thyroid lesions has resulted in unnecessary treatment. An accurate detection of TC at an early stage is highly demanded. We aim to develop an enhanced isobaric labeling-based high-throughput plasma quantitative proteomics to identify biomarkers in a discovery cohort. Selected candidates were tested by enzyme-linked immunosorbent assay (ELISA) in the training cohort and validation cohort. In total, 1063 proteins were quantified, and 129 proteins were differentially expressed between patients and healthy subjects. Serum levels of ISG15 and PLXNB2 were significantly elevated in patients with papillary thyroid cancer (PTC) or thyroid adenoma, compared to healthy subjects (p < 0.001) and patients with nodular goiter (p < 0.001). Receiver operating characteristic (ROC) analysis of combined markers (ISG15 and PLXNB2) significantly distinguished PTC from healthy control (HC) subjects. Similar differentiations were also found between thyroid adenoma and HC subjects. Notably, this combined marker could distinguish stage-I PTC from HC subjects (area under the curve (AUC) = 0.872). Our results revealed that ISG15 and PLXNB2 are independent diagnostic biomarkers for PTC and thyroid adenoma, showing a promising value for the early detection of PTC.

hsa-miR-206b Involves in the Development of Papillary Thyroid Carcinoma via Targeting LMX1B.

Objectives: Papillary thyroid carcinoma (PTC) is the most common endocrine system malignant thyroid cancer, and patients with lymph node metastasis typically exhibit poor prognosis. MicroRNAs (miRNAs) can act as either oncogenes or tumor suppressors in PTC. This study was aimed at using PTC transcriptome data obtained from The Cancer Genome Atlas (TCGA) to identify differentially expressed, survival-related miRNAs and target genes. Methods: We analyzed the TCGA datasets to identify differentially expressed mRNAs/miRNAs in 493 PTC patients with stage I_II group (stages I and II) versus stage III_IV group (stages III and IV) according to TNM staging. The Kaplan-Meier survival analysis, the Cox regression analysis, and the log-rank test were performed to investigate survival-related miRNAs. Results: We identified 36 significantly differentially expressed miRNAs in the stage I_II group versus the stage III_IV group, in which 31 were upregulated and only 5 were downregulated (i.e., hsa-miR-891a-5p, hsa-miR-892a, hsa-miR-888-5p, hsa-miR-891b, and hsa-miR-892b). Additionally, five signature miRNAs (hsa-miR-206, hsa-miR-299-3p, hsa-miR-299-5p, hsa-miR-496, and hsa-miR-509-3-5p) were associated with the overall survival of PTC patients. We also found that LMX1B, whose expression was inversely correlated with hsa-miR-206 expression, was a putative target gene of hsa-miR-206 and LMX1B was likely to serve as a tumor suppressor in PTC. Conclusion: hsa-miR-206b might be involved in promoting TNM staging in PTC via targeting of LMX1B.

TFE3 Regulates the Function of the Autophagy-Lysosome Pathway to Drive the Invasion and Metastasis of Papillary Thyroid Carcinoma.

BACKGROUND: Accumulating evidence shows that autophagy plays a vital role in tumor occurrence, development, and metastasis and even determines tumor prognosis. However, little is known about its role in papillary thyroid carcinoma (PTC) or the potentially oncogenic role of TFE3 in regulating the autophagy-lysosome system. METHODS: Immunohistochemistry and quantitative real-time PCR (qRT-PCR) were used to examine the expression of TFE3, P62/SQSTM1, and LC3 in PTC and paracancerous tissues. TFE3, P62/SQSTM1, LC3, cathepsin L (CTSL), and cathepsin B (CTSB) were evaluated using Western blot analysis. After inducing TFE3 overexpression by plasmid or TFE3 downregulation by small interfering RNA (siRNA) transfection, MTT, wound healing, and cell migration and invasion assays were used to verify the effects on invasion, migration, and the levels of autophagy-lysosome system-related proteins such as P62/SQSTM1, LC3, CTSL, and CTSB. RESULTS: TFE3 was overexpressed in PTC tissues compared with paracancerous tissues. Analysis of the clinicopathological characteristics of PTC patients showed that high TFE3 expression was significantly correlated with lymph node metastasis. TFE3 overexpression in the PTC cell lines KTC-1 and BCPAP promoted proliferation, invasion, and migration, while TFE3 knockdown had the opposite effects. Furthermore, we identified a positive relationship among the expression levels of TFE3, P62/SQSTM1, LC3, CTSL, and CTSB. We found that silencing TFE3 inhibited the expression of P62/SQSTM1, LC3, CTSL, and CTSB in PTC cells. However, TFE3 overexpression had the opposite effects. CONCLUSIONS: The present study provided evidence for the underlying mechanisms by which TFE3 induces autophagy-lysosome system activity in PTC.

T cell exhaustion is associated with the risk of papillary thyroid carcinoma and can be a predictive and sensitive biomarker for diagnosis.

BACKGROUND: The incidence of papillary thyroid carcinoma (PTC) has been steadily increasing over the past decades. Hashimoto's thyroiditis (HT) is the most common autoimmune disease, and is related to the pathogenesis of PTC. Programmed death-1 (PD-1) is currently used for the treatment of PTC, but there are very few studies on the clinical value of PD-1 in the diagnosis and targeted therapy of PTC. METHODS: The expression of T, B, NK cells and PD-1 in the peripheral blood of 132 patients with PTC (PTC group), 48 patients with nodular goiter (NG group) and 63 healthy subjects (HP group) were detected by flow cytometry. The expression of plasma T3, T4, FT3, FT4, TSH, TGAb and TPO was detected by chemiluminescence immunoassay. Among 132 PTC, 49 PTC&HT and 83 PTC&noHT were included. Among 48 NG, 10 NG&HT and 38 NG&noHT were included. The expressions of programmed death- ligand1(PD-L1) in tumor tissues of PTC group and thyroid tissues of NG group, PD-1 and CD3 in tumor infiltration lymphocyte (TIL) were detected by immunohistochemistry. RESULTS: The expression of FT3, TGAb, CD3+PD-1+, CD3+CD4+PD-1+ and CD3+CD8+PD-1+ in PTC and NG was significantly higher than that in the HP group. Moreover, CD3+PD-1+, CD3+CD4+PD-1+ and CD3+CD8+PD-1+ expression had significant differences between the PTC group and the NG group. In addition, the expression of TGAb, TPO, CD3+PD-1+, CD3+CD4+PD-1+ and CD3+CD8+PD-1+ in PTC&HT group was significantly higher than that in the PTC&noHT group. While, the expression of B cells, CD3+PD-1+, CD3+CD4+PD-1+ and CD3+CD8+PD-1+ in PTC&HT group was higher than that in NG&HT group. PD-1 showed a significant correlation with PTC lymph node metastasis. CD3+PD-1+ and CD3+CD4+PD-1+ was higher in N1 stage than in N0 stage. Immunohistochemical results showed that the expression of PD-1, CD3 and PD-L1 in PTC was significantly higher than that in NG. CONCLUSIONS: T cell exhaustion might act as a biomarker for the differential diagnosis of PTC and NG. Patients with PTC&HT have obvious T cell exhaustion and increased expression of PD-1, PD-L1.Targeting the PD-1/PD-L1 pathway could be a new approach to prevent malignant transformation from HT to PTC&HT in the future.

Relationship of oestrogen receptor alpha gene polymorphisms with risk for benign prostatic hyperplasia and prostate cancer in Chinese men.

The relationship of oestrogen receptor with benign prostatic hyperplasia (BPH) and prostate cancer (PC) is not clear at present. This study aimed to investigate the molecular mechanism underlying the occurrence and development of BPH and prostate.Two hundred forty-four PC cases, 260 BPH patients, and 222 healthy men were recruited from Han people in China, and the oestrogen receptor alpha (ESRα) gene polymorphism (rs2234693 [PvuII] and rs9340799 [XbaI]) on intron 1 was determined. The relationship of gene polymorphism with PC and BPH was evaluated with Logistic regression, and the linkage disequilibrium and haplotyping were assessed with SHEsis software.The risk for PC in BPH patients with PvuII C allele was higher (OR = 1.437, 95% CI: 1.110-1.859), but the differentiation degree of cancer cells was relatively better in PC patients with PvuII C allele (OR = 0.419, 95% CI: 0.285-0.616), and most of them are circumscribed (OR = 0.706, 95% CI: 0.485-1.02). There was significant linkage disequilibrium between PvuII and XbaI. The genotype TTAG not only induced BPH (OR = 6.260, 95% CI: 1.407-27.852), but increased the risk for PC (OR = 6.696, 95% CI: 1.504-29.801). However, the genotype TTAG in BPH patients had no relationship with the risk for PC (P > 0.05). Furthermore, men with haplotype TG were more likely to suffer PC (OR = 9.168, 95% CI: 2.393-35.119), but men with haplotype TA and enlarged prostate had a low risk for PC (OR = 0.708, 95% CI: 0.551-0.912).These results show the relationship between ESRα gene polymorphism and susceptibility to PC and BPH in Chinese men, and the ethnic and regional difference as well.

Celastrol may have an anti-atherosclerosis effect in a rabbit experimental carotid atherosclerosis model.

BACKGROUND: Celastrol may have an anti-atherosclerosis effect. This study aimed to investigate if celastrol had an anti-AS effect using a rabbit experimental carotid atherosclerosis model. METHODS: Forty male Japanese white rabbits were divided into the sham group (normal diet), the model group (high fat diet), the group treated with celastrol (high fat diet) and the group treated with atorvastatin (high fat diet) randomly. The rabbits fed a high fat diet underwent balloon injury of the right common carotid artery and were treated with dimethyl sulfoxide (DMSO) (the model group, 3.5 ml/kg/d), celastrol and its dissolvent DMSO (the celastrol group, 1 mg/kg/d and 3.5 ml/kg/d) and atorvastatin and its dissolvent DMSO (the atorvastatin group, 2.5 mg/kg/d and 3.5 ml/kg/d) for 12 weeks by gavage. RESULTS: The ratio of the plaque area and the arterial wall cross-section area in the celastrol group was significantly less than the model group (P < 0.001), and there was no significant difference compared with the atorvastatin group. The serum level of LDL-C of the celastrol group was significantly lower than the model group (P = 0.014), and there was no significant difference compared with the atorvastatin group. The expression of VEGF in the celastrol group was significantly less compared with the model group (P = 0.014), whereas the expression of VEGF in the atorvastatin group and the model group showed no significant differences. CONCLUSION: Our findings suggest that celastrol effectively reduced the plaque ratio, decreased the serum levels of LDL and downregulated the expression of VEGF, suggesting an anti-AS effect of celastrol.

The relationship between polymorphisms in the promoter region of Tim-3 and unexplained recurrent spontaneous abortion in Han Chinese women.

BACKGROUND: Recurrent spontaneous abortion (RSA) refers to 2 or more consecutive pregnancy losses, and RSA with unknown causes is called unexplained recurrent spontaneous abortion (URSA). Tim-3, a subtype of the T-cell immunoglobulin domain and mucin domain (Tim) protein family, might be an important regulatory molecule that plays a pivotal role in URSA, which might be triggered mostly by Th1/Th2 immune deviation. To understand the etiology and pathogenesis of URSA in Han Chinese women, we investigated the association between polymorphisms of rs10053538 and rs10515746 in the promoter of Tim-3 and the risk of URSA in Han Chinese women. METHODS: One hundred and forty-eight women with RSA resulting in still birth were enrolled in the URSA group. We performed tests to rule out congenital reproductive system malformation, reproductive system tumor, endocrine dyscrasia, and chromosome abnormalities. One hundred and fifty-three women with normal pregnancy leading to live birth were selected at random to comprise the control group. All women included in this study were genetically unrelated Han Chinese women. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and allele-specific polymerase chain reaction (AS-PCR) were used to determine polymorphisms of rs10053538 and rs10515746, respectively, in all subjects. PCR products were chosen at random for sequencing. RESULTS: No significant statistical difference was found between the distribution frequency of the GT + TT genotype and T allele on the rs10053538 locus in the URSA group or the control group (10.1% vs. 11.8%, Chi(2) = 0.205, P = 0.651; 5.1% vs. 6.5%, Chi(2) = 0.592, P = 0.441; respectively). Neither was there a significant difference between the distribution frequency of the GT + TT genotype and T allele on the rs10515746 locus in the groups (6.8% vs. 3.9%, Chi(2)1.201, P = 0.273; 3.4% vs. 2.0%, Chi(2) = 1.169, P = 0.280; respectively). CONCLUSIONS: The present study suggested that these polymorphisms of rs10053538 or rs10515746 in the Tim-3 promoter may not be associated with URSA in Han Chinese women.