Clinical value of multi-gene testing in distinguishing benign and malignant thyroid nodules.

BACKGROUND: The newly released 2022 WHO Classification of Neuroendocrine Neoplasms (version 5) and a recent update on thyroid tumor classifications have emphasized genetic testing to an unprecedented level. Fine needle aspiration (FNA) has been widely applied for the preoperative diagnosis of thyroid nodules. However, it is limited mainly to testing for a single gene-BRAFV600E, whereas multi-gene testing data are scarce, especially in the Asian population. This study aimed to explore the clinical value of multi-gene testing in the differential diagnosis of benign and malignant thyroid nodules based on the 2023 Bethesda System for Reporting Thyroid Cytopathology (BSRTC). METHODS: A total of 615 thyroid nodules underwent ultrasound-guided fine-needle aspiration cytology (FNAC) were collected from Sir Run Run Shaw Hospital, Zhejiang University School of Medicine. The next-generation sequencing platform was applied for multi-gene testing. A panel of well-recognized commonly mutated genes in thyroid cancer were analyzed, including BRAFV600E, KRAS, NRAS, HRAS, TERT, TP53, PAX8/PPARG, CCDC6/ RET and NCOA4/ RET. RESULTS: Gene mutations were identified in 324 nodules (52.7%), with BRAFV600E being the most prevalent driver gene alteration observed in this cohort (233/324; 79.1%), followed by RAS (77/324, 23.8%). The overall malignancy rate of gene mutations was 89.7% in our cohort, of which the lymph node metastasis rate was 45.3%. The combination of multi-gene testing and cytology resulted in 89.3% sensitivity, 95.2% specificity, 98.9% positive predictive value, 64.5% negative predictive value and 90.3% accuracy, which were significantly higher than those from mere cytology (sensitivity 68.6%, specificity 87.5%, positive predictive value 95.9%, negative predictive value 39.8%, accuracy 72.2%). CONCLUSIONS: Multi-gene testing could substantially enhance the detection rate of malignant thyroid nodules and protect patients with benign nodules from unnecessary surgeries. Multi-gene testing provides a valuable reference for individualized preoperative decision-making, which may serve as a crucial method for postoperative treatment and prognosis assessment.

Comparison of S-Detect and thyroid imaging reporting and data system classifications in the diagnosis of cytologically indeterminate thyroid nodules.

Purpose: The aim of this study was to investigate the value of S-Detect for predicting the malignant risk of cytologically indeterminate thyroid nodules (CITNs). Methods: The preoperative prediction of 159 CITNs (Bethesda III, IV and V) were performed using S-Detect, Thyroid Imaging Reporting and Data System of American College of Radiology (ACR TI-RADS) and Chinese TI-RADS (C-TIRADS). First, Linear-by-Linear Association test and chi-square test were used to analyze the malignant risk of CITNs. McNemar's test and receiver operating characteristic curve were used to compare the diagnostic efficacy of S-Detect and the two TI-RADS classifications for CITNs. In addition, the McNemar's test was used to compare the diagnostic accuracy of the above three methods for different pathological types of nodules. Results: The maximum diameter of the benign nodules was significantly larger than that of malignant nodules [0.88(0.57-1.42) vs 0.57(0.46-0.81), P=0.002]. The risk of malignant CITNs in Bethesda system and the two TI-RADS classifications increased with grade (all P for trend<0.001). In all the enrolled CITNs, the diagnostic results of S-Detect were significantly different from those of ACR TI-RADS and C-TIRADS, respectively (P=0.021 and P=0.007). The sensitivity and accuracy of S-Detect [95.9%(90.1%-98.5%) and 88.1%(81.7%-92.5%)] were higher than those of ACR TI-RADS [87.6%(80.1%-92.7%) and 81.8%(74.7%-87.3%)] (P=0.006 and P=0.021) and C-TIRADS [84.3%(76.3%-90.0%) and 78.6%(71.3%-84.5%)] (P=0.001 and P=0.001). Moreover, the negative predictive value and the area under curve value of S-Detect [82.8% (63.5%-93.5%) and 0.795%(0.724%-0.855%)] was higher than that of C-TIRADS [54.8%(38.8%-69.8%) and 0.724%(0.648%-0.792%] (P=0.024 and P=0.035). However, the specificity and positive predictive value of S-Detect were similar to those of ACR TI-RADS (P=1.000 and P=0.154) and C-TIRADS (P=1.000 and P=0.072). There was no significant difference in all the evaluated indicators between ACR TI-RADS and C-TIRADS (all P>0.05). The diagnostic accuracy of S-Detect (97.4%) for papillary thyroid carcinoma (PTC) was higher than that of ACR TI-RADS (90.4%) and C-TIRADS (87.8%) (P=0.021 and P=0.003). Conclusion: The diagnostic performance of S-Detect in differentiating CITNs was similar to ACR TI-RADS and superior to C-TIRADS, especially for PTC.

The combination of BRAFV600E mutation and Chinese Thyroid Imaging Reporting and Data System is helpful in the management of AUS/FLUS thyroid nodules.

PURPOSE: To explore the utility of the BRAFV600E mutation in combination with the Chinese Thyroid Imaging Reporting and Data System (C-TIRADS) in the management of atypia of undetermined significance/follicular lesion of undetermined significance (AUS/FLUS) thyroid nodule (TN). METHODS: 138 AUS/FLUS TNs in 129 patients were included. Each TN underwent preoperative BRAFV600E mutation analysis and was classified using the C-TIRADS. Histopathologic diagnosis served as reference standard. RESULTS: 46 benign TNs and 92 malignant TNs were identified. The C-TIRADS 4C and 5 (OR = 10.409, P = 0.000), BRAFV600E mutation (OR = 36.493, P = 0.000) were independent predictors of malignant nodules. There were significant differences in malignancy rate among the different C-TIRADS TNs (P = 0.000), and these TNs with higher C-TIRADS were associated with increased malignancy rate (P for trend = 0.000). The rate of the nodule with BRAFV600E mutation increased with the increase of C-TIRADS (P for trend = 0.001). For AUS/FLUS TNs without BRAFV600E mutation, the malignancy rates of the C-TIRADS 3, 4A, 4B, 4C, and 5 were 0%, 21.4%, 20.8%, 70.8%, and 100%, respectively (P = 0.000), and the malignancy rate increased from C-TIRADS 3 to C-TIRADS 5 (P for trend = 0.000). C-TIRADS and BRAFV600E mutation had similar diagnostic efficacy (P > 0.05), and the sensitivity, negative predictive value, and accuracy of the combination were significantly higher than BRAFV600E gene or C-TIRADS alone (P < 0.05). CONCLUSIONS: C-TIRADS can effectively provide risk stratification for AUS/FLUS nodules. The combination is helpful in selecting appropriate management for AUS/FLUS patients.

BRAF V600E mutation and the Bethesda System for Reporting Thyroid Cytopathology of fine-needle aspiration biopsy for distinguishing benign from malignant thyroid nodules.

BACKGROUND: The Bethesda System for Reporting Thyroid Cytopathology (TBSRTC) predicts the risk of malignancy for the different categories of the ultrasound-guided fine-needle aspiration biopsy (FNAB). The objective of this study is to investigate the efficiencies of the v-raf murine sarcoma viral oncogene homolog B1 (BRAF) V600E mutation test and the TBSRTC categories in distinguishing between benign and malignant thyroid nodules. METHODS: In this study, 362 ultrasound-guided fine-needle aspiration (FNA) samples from 344 patients aged from 17 to 76 years old were retrospectively reviewed. The patients were classified into six groups (I-VI) according to the TBSRTC system. The amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) was used to evaluate the BRAF V600E mutation level in total 362 samples. Among of the 344 patients, 128 patients (131 thyroid nodules) who underwent surgeries were followed by histopathological examination. The predictive values of the BRAF V600E mutation test and TBSRTC categories were evaluated in these 131 thyroid nodules. RESULTS: The median ages of the patients in the TBSRTC IV-VI group were smaller than those in the TBSRTC I-III groups. The proportion of nodules over 1 cm was larger than it in the TBSRTC IV group compared to the other groups. Significant differences in BRAF V600E mutation were observed (P < .001) among these six groups. The sensitivity (89.57%) for the detection of malignant thyroid nodules, negative predictive value (NPV; 45.45%) for the detection of benign nodules, and accuracy (86.26%) for distinguishing between benign and malignant thyroid nodules increased by combining the BRAF V600E mutation test and TBSRTC system when compared with the BRAF V600E mutation test and TBSRTC system respectively. The BRAF V600E mutation test alone demonstrated the increased positive predictive value (PPV; 98.91%) and specificity (93.75%) for the detection of malignant thyroid nodules compared to the TBSRTC method (alone or in combination with the BRAF V600E method). CONCLUSION: In summary, significant differences in age, nodule diameter, and BRAF V600E mutation were noted among the six categories of the TBSRTC system. The combination of the BRAF V600E mutation test and TBSRTC system demonstrated increases in the NPV, sensitivity, and accuracy, while the BRAF V600E method proved superiority to the TBSRTC system with regard to the PPV and specificity.

Role of preoperative washout fluid thyroglobulin from fine-needle aspirates in detecting lymph node metastases of papillary thyroid carcinoma and its influential factors / 中国肿瘤临床

Objective: To investigate the diagnostic value and influential factors of washout fluid thyroglobulin collected during fine-needle aspiration (FNA-Tg) in detecting lymph node metastases of papillary thyroid carcinoma (PTC) before thyroidectomy. Methods:We retrospectively analyzed 131 patients diagnosed with PTC based on histopathology. They presented with suspicious enlarged cervi-cal lymph nodes and underwent high-frequency ultrasound-guided FNA before the surgery. FNA and FNA-Tg were performed simulta-neously. All the related data were collected. In order to obtain the best cut-off value, the FNA-Tg receiver-operating characteristic curve was generated. The cytopathology and postoperative pathologic results, as well as the ultrasound images during the follow-up, were considered the gold standard. The diagnostic performance of each method (FNA, FNA-Tg, and FNA+FNA-Tg) were compared. Ad-ditionally, some suspicious influential factors such as the anatomical location of lymph nodes and associated laboratory indexes were also analyzed for the diagnostic accuracy of FNA-Tg. Results: The best cut-off value of FNA-Tg in our study was 1.295 ng/mL. The diag-nostic performance of the combined method was the best when compared with other methods, with a sensitivity of 96.4% and speci-ficity of 99.2%. Additionally, FNA-Tg was much more accurate when used in diagnosis of lateral cervical lymph nodes. Among all the as-sociated laboratory indexes, the level of serum Tg (sTg) was an independent predictive factor for an FNA-Tg level above 1.295 ng/mL (odds ratio=1.018). Conclusions: FNA-Tg is a useful tool in the identification of metastatic cervical lymph nodes preoperatively, espe-cially for lateral cervical lymph nodes. In addition, 1.295 ng/mL could be one of the reference standards of the FNA-Tg cut-off value. When the sTg level is high, we should interpret the FNA-Tg results cautiously.

[The value of thyroglobulin measurement in fine-needle aspiration for diagnosis of suspicious lymph nodes in patients with thyroid carcinoma after thyroidectomy].

OBJECTIVE: To investigate the value of thyroglobulin measurement in ultrasound guided fine-needle aspiration(FNA-Tg)for detecting neck node metastasis in patients with papillary thyroid carcinoma(PTC)after thyroidectomy. METHODS: A total of 128 suspicious metastatic lymph nodes in 112 patients were retrospectively analyzed. Postoperative pathologic results were taken as the gold standard. The values of FNA and FNA-Tg combined FNA in the diagnosis of metastatic lymph nodes were compared with different ultrasonic features. RESULTS: The sensitivity, specificity and accuracy of single FNA for the diagnosis of neck node metastasis were 67.5%, 98.0% and 79.7% respectively, and those of FNA-Tg combined FNA were 87.0%, 100.0% and 92.2% respectively. Compared with single FNA, the sensitivity and accuracy of FNA-Tg combined FNA were increased significantly(χ(2)=8.319, P=0.004; χ(2)=8.275, P=0.004). When the ultrasonographic characteristics met any one of five indicators for neck node metastasis, the sensitivity, specificity and accuracy of single FNA for the diagnosis of neck node metastasis were 38.1%, 95.7% and 68.2% respectively, and those of FNA-Tg combined FNA were 71.0%, 100.0% and 86.4% respectively. Compared with single FNA, the sensitivity and accuracy of FNA-Tg combined FNA were increased significantly(χ(2)=4.709, P=0.030; χ(2)=4.141, P=0.042). When the ultrasonographic characteristics met any two of five indicators for neck node metastasis, the sensitivity, specificity and accuracy of single FNA for the diagnosis of neck node metastasis were 55.0%, 100.0% and 73.5% respectively, and those of FNA-Tg combined FNA were 90.0%, 100.0% and 94.1% respectively. Compared with single FNA, the sensitivity and accuracy of FNA-Tg combined FNA were increased significantly(χ(2)=6.140, P=0.013; χ(2)=5.314, P=0.021). The ultrasonographic characteristics met any three, four or five of five indicators or did not meet any of the indicators, there was no significant difference in the value of the diagnosis of metastatic lymph nodes between single FNA and FNA-Tg combined FNA(P>0.05). CONCLUSIONS: The combination of FNA with FNA-Tg could be helpful in diagnosis of lymph node metastasis. When the suspicious lymph nodes had one or two ultrasound characteristics for neck node metastasis, FNA-Tg could raise the sensitivity and accuracy of FNA, and FNA-Tg could not significantly improve in the diagnosis of FNA when presenting with no or with more than 2 ultrasonographic features.

Rutaecarpine suppresses atherosclerosis in ApoE-/- mice through upregulating ABCA1 and SR-BI within RCT.

ABCA1 and scavenger receptor class B type I (SR-BI)/CD36 and lysosomal integral membrane protein II analogous 1 (CLA-1) are the key transporter and receptor in reverse cholesterol transport (RCT). Increasing the expression level of ABCA1 and SR-BI/CLA-1 is antiatherogenic. The aim of the study was to find novel antiatherosclerotic agents upregulating expression of ABCA1 and SR-BI/CLA-1 from natural compounds. Using the ABCA1p-LUC and CLA-1p-LUC HepG2 cell lines, we found that rutaecarpine (RUT) triggered promoters of ABCA1 and CLA-1 genes. RUT increased ABCA1 and SR-BI/CLA-1 expression in vitro related to liver X receptor alpha and liver X receptor beta. RUT induced cholesterol efflux in RAW264.7 cells. ApoE-deficient (ApoE(-/-)) mice treated with RUT for 8 weeks showed ∼68.43, 70.23, and 85.56% less en face lesions for RUT (L), RUT (M), and RUT (H) groups, respectively, compared with the model group. Mouse macrophage-specific antibody and filipin staining indicated that RUT attenuated macrophages and cholesterol accumulations in atherosclerotic lesions, respectively. Additionally, ABCA1 and SR-BI expression was highly induced by RUT in livers of ApoE(-/-) mice. Meanwhile, RUT treatment significantly increased the fecal (3)H-cholesterol excretion, which demonstrated that RUT could promote RCT in vivo. RUT was identified to be a candidate that protected ApoE(-/-) mice from developing atherosclerosis through preferentially promoting activities of ABCA1 and SR-BI within RCT.

The role of slingshot-1L (SSH1L) in the differentiation of human bone marrow mesenchymal stem cells into cardiomyocyte-like cells.

Adult cardiomyocytes (CMs) have very limited capacity to regenerate. Therefore, there is a great interest in developing strategies to treat infarcted CMs that are able to regenerate cardiac tissue and promote revascularization of infarcted zones in the heart. Recently, stem cell transplantation has been proposed to replace infarcted CMs and to restore the function of the affected tissue. This area of research has become very active in recent years due to the huge clinical need to improve the efficacy of currently available therapies. Slingshot (SSH) is a family of protein phosphatases, which can specifically dephosphorylate and reactivate cofilin and inhibit the polymerization of actin filaments and actively involved in cytoskeleton rearrangement. In this study, we found that SSH1L promoted morphology changes of microfilaments during differentiation but was inhibited by the inhibitors of actin polymerization such as cytochalasin D. Overexpression of SSH1L could promote cardiac-specific protein and genes expression. 5-Aza can induce the differentiation of hMSCs into cardiomyocyte-like cells in vitro. We also observed that SSH1L efficiently promotes hMSCs differentiation into cardiomyocyte-like cells through regulation and rearrangement of cytoskeleton. Our work provides evidence that supports the positive role of SSH1L in the mechanism of stem cell differentiation into cardiomyocyte-like cells.