Detection of BRAF V600E in Fine-Needle Aspiration Samples of Thyroid Nodules by Droplet Digital PCR.

Background: BRAF exon 15 p.V600E (BRAF V600E) mutation has been established as an important molecular marker for papillary thyroid carcinoma diagnosis by ultrasound-guided fine-needle aspiration biopsy (FNAB). Sanger sequencing is the gold standard for detecting BRAF V600E mutations but fails to identify low-frequency mutations. However, droplet digital PCR (ddPCR) is a popular new method for detecting low-frequency mutations. Here, we compare the efficiency of droplet digital PCR (ddPCR) and Sanger sequencing for detection of the BRAF V600E mutation in thyroid fine-needle aspiration (FNA) samples. Methods: Thyroid fine-needle aspiration samples from 278 patients with 310 thyroid nodules were collected. Sanger sequencing and ddPCR were conducted to detect the BRAF V600E mutation. Results: The BRAF V600E mutation was found in 94 nodules (30.32%) by ddPCR and 40 nodules (12.90%) by Sanger sequencing in 310 FNA samples. A total of 119 nodules were confirmed PTC by postsurgical pathology. Among which the BRAF mutation was found in 80 (67.23%) nodules by ddPCR and 31 (26.05%) by Sanger sequencing. All nodules carrying the mutation detected by Sanger sequencing (SS+) were verified by ddPCR (ddPCR+). Also, all nodules with no mutation detected by ddPCR were interpreted as wild-type by Sanger sequencing (SS-). In addition. Almost all SS+/ddPCR + nodules (95.00%; 38/40) and SS-/ddPCR + nodules (100.00%; 54/54) displayed a BRAF mutation rate of >5% and <15%, respectively, indicating easy misdetection by Sanger sequencing when the mutation rate is between 5 and 15%. Conclusion: ddPCR has higher sensitivity than Sanger sequencing and we propose ddPCR as a supplement to Sanger sequencing in molecular testing of BRAF using FNAB samples.

A five-gene panel refines differential diagnosis of thyroid nodules.

BACKGROUND: Molecular testing for oncogenic mutations in fine-needle aspiration has showed high predictive value in identifying malignant lesions from thyroid nodules with indeterminate cytology. METHODS: To figure out an efficient and economical gene panel for most medical institutions in China, we designed a five-gene panel including BRAF/NRAS/KRAS/HRAS/TERT genes and conducted a retrospective study to evaluate the role of this five-gene diagnostic panel in differential diagnosis of thyroid nodules. RESULTS: A total of 665 patients with 695 thyroid nodules were investigated in the current study. The fine-needle aspiration biopsy and surgically separated thyroid tissue specimens were harvested to test BRAF, TERT, NRAS, KRAS, and HRAS mutations. We identified 261 mutations in 665 patients, including 177 V600E mutations in BRAF. Three hundred and sixty-nine patients who underwent thyroid surgery after completion of the initial clinical and cytological evaluation were enrolled in the final analysis. The diagnostic sensitivity, specificity, and accuracy of the combination of FNAB cytology and five-gene detection were 74.7%, 93.8%, and 84.8%, respectively. BRAF V600E and five-gene panel could recognize 46.4% and 53.6% of papillary thyroid carcinoma in the patients with cytologically indeterminate nodules. CONCLUSION: The five-gene panel can effectively improve the sensitivity, negative predictive value, and accuracy of fine-needle aspiration biopsy cytology, especially in the patients with cytologically indeterminate nodules.

Relationship between Free Thyroxine and Islet Beta-cell Function in Euthyroid Subjects.

Thyroid hormones have a specific effect on glucose-induced insulin secretion from the pancreas. We aimed to investigate the association between euthyroid hormones and islet beta-cell function in general population and non-treated type 2 diabetes mellitus (T2DM) patients. A total of 5089 euthyroid participants (including 4601 general population and 488 non-treated T2DM patients) were identified from a cross-sectional survey on the prevalence of metabolic diseases and risk factors in East China from February 2014 to June 2016. Anthropometric indices, biochemical parameters, and thyroid hormones were measured. Compared with general population, non-treated T2DM patients exhibited higher total thyroxine (TT4) and free thyroxine (FT4) levels but lower ratio of free triiodothyronine (T3):T4 (P<0.01). HOMA-ß had prominently negative correlation with FT4 and positive relationship with free T3:T4 in both groups even after adjusting for age, body mass index (BMI) and smoking. When analyzed by quartiles of FT4 or free T3:T4, there were significantly decreased trend of HOMA-ß going with the higher FT4 and lower free T3:T4 in both groups. Linear regression analysis showed that FT4 but not FT3 and free T3:T4 was negatively associated with HOMA-ß no matter in general population or T2DM patients, which was independent of age, BMI, smoking, hypertension and lipid profiles. FT4 is independently and negatively associated with islet beta-cell function in euthyroid subjects. Thyroid hormone even in reference range could play an important role in the function of pancreatic islets.

Low vitamin D levels and non-alcoholic fatty liver disease, evidence for their independent association in men in East China: a cross-sectional study (Survey on Prevalence in East China for Metabolic Diseases and Risk Factors (SPECT-China)).

Recent studies have suggested an association between vitamin D and non-alcoholic fatty liver disease (NAFLD); however, some results are subject to debate. This study was carried out to evaluate the correlation between NAFLD and vitamin D in men and women in East China. The data were obtained from a cross-sectional study that focused on the health and metabolic status of adults in sixteen areas of East China. According to ultrasonic assessments, the patients were divided into normal and NAFLD groups. Demographic characteristics and biochemical measurements were obtained. Binary logistic regression analysis was used to explore the association. In total, 5066 subjects were enrolled, and 2193 (43·3 %) were diagnosed with NAFLD; 84·56 % of the subjects showed vitamin D deficiency. Subjects with high vitamin D levels had a lower prevalence of NAFLD, particularly male subjects. Within the highest quartile of vitamin D levels, the prevalence of NAFLD was 40·8 %, whereas the lowest quartile of vitamin D levels showed a prevalence of 62·2 %, which was unchanged in women across the vitamin D levels. Binary logistic analysis showed that decreased vitamin D levels were associated with an increased risk of NAFLD (OR 1·54; 95 % CI 1·26, 1·88). This study suggests that vitamin D levels are significantly associated with NAFLD and that vitamin D acts as an independent factor for NAFLD prevalence, particularly in males in East China. Vitamin D interventional treatment might be a new target for controlling NAFLD; elucidating the mechanism requires further research.

Characterization of Chondrogenic Gene Expression and Cartilage Phenotype Differentiation in Human Breast Adipose-Derived Stem Cells Promoted by Ginsenoside Rg1 In Vitro.

BACKGROUND/AIMS: Investigating and understanding chondrogenic gene expression during the differentiation of human breast adipose-derived stem cells (HBASCs) into chondrogenic cells is a prerequisite for the application of this approach for cartilage repair and regeneration. In this study, we aim to characterize HBASCs and to examine chondrogenic gene expression in chondrogenic inductive culture medium containing ginsenoside Rg1. METHODS: Human breast adipose-derived stem cells at passage 3 were evaluated based on specific cell markers and their multilineage differentiation capacity. Cultured HBASCs were treated either with basic chondrogenic inductive conditioned medium alone (group A, control) or with basic chondrogenic inductive medium plus 10 µg/ml (group B), 50 µg/ml (group C), or 100µg/ml ginsenoside Rg1 (group D). Cell proliferation was assessed using the CCK-8 assay for a period of 9 days. Two weeks after induction, the expression of chondrogenic genes (collagen type II, collagen type XI, ACP, COMP and ELASTIN) was determined using real-time PCR in all groups. RESULTS: The different concentrations of ginsenoside Rg1 that were added to the basic chondrogenic inductive culture medium promoted the proliferation of HBASCs at earlier stages (groups B, C, and D) but resulted in chondrogenic phenotype differentiation and higher mRNA expression of collagen type II (CO-II), collagen type XI (CO-XI), acid phosphatase (ACP), cartilage oligomeric matrix protein (COMP) and ELASTIN compared with the control (group A) at later stages. The results reveal an obvious positive dose-effect relationship between ginsenoside Rg1 and the proliferation and chondrogenic phenotype differentiation of HBASCs in vitro. CONCLUSIONS: Human breast adipose-derived stem cells retain stem cell characteristics after expansion in culture through passage 3 and serve as a feasible source of cells for cartilage regeneration in vitro. Chondrogenesis in HBASCs was found to be prominent after chondrogenic induction in conditions containing ginsenoside Rg1.

Efficacy and side-effects of a semi-individualized Chinese herb mixture "Tiáo Geng Tang" for menopausal syndrome in China.

BACKGROUND: Chinese herbal medicine is an alternative therapy for menopausal problems and is widely practiced in China and many other Asian countries. However, efficacies and side-effects are rarely assessed according to the standards of evidence-based medicine. PATIENTS AND METHODS: This is a prospective observatory study following efficacy and side-effects of a semi-individualized Chinese herbal mixture "Tiáo Geng Tang (TGT)" in 30 patients for 3 months. Another group of 30 patients receiving hormone therapy with tibolone was included as a positive comparison. Common questionnaire-based measuring instruments were: modified Kupperman index, menopause rating scale, life quality and Chinese medical symptom scale (CMSS). Follicle-stimulating hormone (FSH), luteinizing hormone (LH) and estradiol (E2) were determined before and three months after the treatments. RESULTS: Significant improvement was seen in overall scores of all the four measurements in both groups. For some symptoms, including dry mouth, tinnitus, poor appetite and constipation, TGT was more effective than tibolone. For psychosocial and sexual sub-scales of life quality, tibolone has a slightly higher remedy rate than TGT. TGT lowered FSH and LH significantly, as tibolone did, but elevated E2 significantly less than tibolone. Various adverse events, including body weight increase, abdomen discomfort, nausea/vomiting, emotional instability, pressure in breasts and dizziness, were reported by patients treated with tibolone, whereas only diarrhea was observed in two patients treated with TGT. CONCLUSION: TGT alleviates menopausal symptoms with similar efficacy as tibolone but has fewer side effects.

[Comprehensive traditional Chinese medicine intervention for perimenopausal syndrome in women: a community study].

BACKGROUND: As perimenopausal syndrome is a particularly disturbing condition to the patient, it is practical and necessary to establish a program of comprehensive traditional Chinese medicine therapy for women with perimenopausal syndrome. OBJECTIVE: To observe the therapeutic effects of a comprehensive traditional Chinese medicine therapy for women with perimenopausal syndrome in the community. DESIGN, SETTING, PARTICIPANTS AND INTERVENTIONS: Women with perimenopausal syndrome who met the inclusion criteria in 3 communities of Shanghai were selected for this study. Comprehensive traditional Chinese medicine therapy, including administration of Chinese herbs, auricular point therapy, psychological counseling and the practice of Tai Chi, was applied for these women. MAIN OUTCOME MEASURES: The modified Kupperman index was measured once every 4 weeks for 12 weeks; the indexes of menopause-specific quality of life (MENQOL) questionnaire were measured before and after the treatment. RESULTS: There was no significant difference in 4 groups of negative conversion ratio of the modified Kupperman index at the 4th, 8th and 12th week, respectively. The indexes of the MENQOL questionnaire showed significant differences in 4 groups before and after the treatment. The overall efficacy rate of the comprehensive traditional Chinese medicine therapy in women with perimenopausal syndrome achieved 97.84% at the 8th and the 12th week. CONCLUSION: The comprehensive traditional Chinese medicine therapy can achieve significant improvements in women with perimenopausal syndrome who exhibit both physical and psychological symptoms. With such significant clinical effects, the comprehensive traditional Chinese medicine therapy should be promoted in the community.