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Detecting BRAF mutation by pyrosequencing is more sensitive, faster, and less expensive than direct DNA sequencing and is proposed as an adjunct diagnostic tool in evaluating thyroid nodules of indeterminate cytology.

Mucosal inflammation is characterized by neutrophil and mononuclear cell infiltration. This study aimed to determine the gastric and duodenal microbiota associated with histological, endoscopic, and symptomatic gastritis. Dyspeptic adults who presented for evaluation were included. Subjects with either comorbidities or recent drug intake were excluded. Three endoscopic biopsies were obtained from the antrum, body, and duodenum. Next-generation sequencing for 16S ribosomal RNA V1–V2 hypervariable regions was performed. The correlation between the composition of microbiota and the degree of inflammatory cell infiltration, endoscopic findings, and Patient Assessment of Gastrointestinal Disorders Symptom Severity Index (PAGI-SYM) score was analyzed. In 98 included subjects, microbial communities in the antrum and body showed Bray–Curtis similarity; however, those in the duodenum showed dissimilarity. Histological and endoscopic gastritis was associated with the abundance of Helicobacter pylori and that of commensal bacteria in the stomach. The abundances of Variovorax paradoxus and Porphyromonas gingivalis were correlated with histological gastritis, but not with endoscopic or symptomatic gastritis. The total PAGI-SYM score showed a stronger correlation with the duodenal microbiota (Prevotella nanceiensis and Alloprevotella rava) than with the gastric microbiota (H. pylori, Neisseria elongate, and Corynebacterium segmentosum). Different correlations of the gastric and duodenal microbiota with histological, endoscopic, and symptomatic gastritis were observed for the first time at the species level. H. pylori-negative gastritis is not associated with endoscopic or symptomatic gastritis. Only H. pylori-induced endoscopic gastritis requires gastric cancer surveillance. Owing to the weak correlation with H. pylori, symptomatic gastritis should be assessed separately from histological and endoscopic gastritis.

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Abstract. Direct sequencing is the standard method for the detection of epidermal growth factor receptor (EGFR) mutations in lung cancer, however, its relatively low sensitivity limits its clinical use. Pyrosequencing is a bioluminometric, real-time non-electrophoretic DNA sequencing technique with a number of advantages compared with direct sequencing, including higher sensitivity, speed, automation and costeffectiveness. Clinical specimens from 202 lung cancer patients were analyzed for EGFR mutations in exons 18, 19, 20 and 21 using the pyrosequencing method following genomic DNA extraction from paraffin-embedded tissue specimens. All clinical data and tumor specimens were obtained from the Konkuk University Hospital (Korea) between July 2006 and December 2008. The results and clinical responses to EGFR-tyrosine kinase inhibitors (TKIs) were compared. Overall, EGFR mutation-positive rate was 26.7% (54/202). Activating EGFR mutations were observed more frequently in female (52.1 vs. 13.0%), non-smoking (47.8 vs. 15.8%) and adenocarcinoma (35.2 vs. 5.2%) patients. However, significant numbers of EGFR mutation-positive patients were identified as male, former or current smokers and non-adenocarcinoma patients. The combinations of favorable clinicopathological factors, including female, non-smoking and adenocarcinoma, were not identified to significantly increase the positive EGFR mutation rate (female, 52.1%; female and non-smoker, 52.6%; female, non-smoker and adenocarcinoma, 51.9%). The present findings indicate that EGFR mutation analysis is a highly useful method for the prediction of response to EGFR-TKI and the use of favorable clinicopathological factors to perform this analysis is not suitable. Exon 19 deletion was the most common mutation (63.6%) and exon 21 L858R substitution was measured at 32.7%. The exon 20 T790M mutation was identified in 1 patient prior to EGFR-TKI treatment. EGFR mutation status is associated with response to EGFR-TKI and the overall response rate in patients who have the activating EGFR mutation was 82.4 vs. 5.9% in patients with a wild-type EGFR. The present study demonstrates that EGFR mutations analyzed by the pyrosequencing method are well correlated with clinicopathological parameters and that this method may be useful in the clinical practice.

Follicular variant of papillary thyroid carcinoma (FVPTC), particularly the encapsulated subtype, often causes a diagnostic dilemma. We reconfirmed the molecular profiles in a large number of FVPTCs and investigated the efficacy of the preoperative mutational analysis in indeterminate thyroid nodules. BRAF V600E/K601E and RAS mutational analysis was performed on 187 FVPTCs. Of these, 132 (70.6%) had a point mutation in one of the BRAF V600E (n = 57), BRAF K601E (n = 11), or RAS (n = 64) genes. All mutations were mutually exclusive. The most common RAS mutations were at NRAS codon 61. FNA aspirates from 564 indeterminate nodules were prospectively tested for BRAF and RAS mutation and the surgical outcome was correlated with the mutational status. Fifty-seven and 47 cases were positive for BRAF and RAS mutation, respectively. Twenty-seven RAS-positive patients underwent surgery and all except one patient had FVPTC. The PPV and accuracy of RAS mutational analysis for predicting FVPTC were 96% and 84%, respectively. BRAF or RAS mutations were present in more than two-thirds of FVPTCs and these were mutually exclusive. BRAF mutational analysis followed by N, H, and KRAS codon 61 mutational analysis in indeterminate thyroid nodules would streamline the management of patients with malignancies, mostly FVPTC.

BackgroundAlbuminuria is generally accepted as a sensitive marker of diabetic nephropathy but has limitations in predicting its progression. The aim of this study was to evaluate the use of nonalbumin proteinuria in addition to albuminuria for predicting the progression of type 2 diabetic nephropathy.MethodsIn this retrospective observational study, the urine albumin-to-creatinine ratio (ACR) and the nonalbumin protein-to-creatinine ratio (NAPCR) were measured in 325 patients with type 2 diabetes and estimated glomerular filtration rates (eGFR) ≥30 mL/min/1.73 m2. The patients were divided into four groups based on the cutoff points for the urinary ACR (30 mg/g) and NAPCR (120 mg/g). The renal outcomes were chronic kidney disease (CKD) progression and accelerated eGFR decline.ResultsDuring the 4.3-year follow-up period, 25 (7.7%) patients showed CKD progression and 69 (21.2%) patients showed accelerated eGFR decline. After adjusting for nine clinical parameters, the group with a NAPCR greater than 120 mg/g exhibited higher cumulative incidences of CKD progression (hazard ratio 6.84; P = 0.001) and accelerated eGFR decline (hazard ratio 1.95; P = 0.011) than the group with a NAPCR < 120 mg/g. In patients with normoalbuminuria, the group with NAPCR levels greater than 120 mg/g also exhibited a higher cumulative incidence than that with NAPCR levels <120 mg/g of CKD progression (hazard ratio 21.82; P = 0.005). The addition of NAPCR to ACR improved the model fit for CKD progression and accelerated eGFR decline.ConclusionNonalbumin proteinuria showed additional value over and above that of albuminuria for predicting the progression of CKD in patients with type 2 diabetes.

Primary squamous cell carcinoma of the thyroid (SCC-T) is extremely rare. Its clinical presentation is similar to that of anaplastic carcinoma. Metastasis or extension from the head and neck area should be ruled out, as patients with SCC-T have a poorer prognosis than patients who have a thyroid extension from an adjacent tumor. An 87-year-old man presented with a longstanding painless mass in the right thyroid and had experienced 2 months of pain upon swallowing. A right lobectomy was performed with resection of thyroid cartilage, cricoid cartilage, a portion of the first to third tracheal ring and the right neck lymph node. A histological examination revealed pure SCC. The tumor cells showed diffuse immunoreactivity to CK5/6, CK19 and p63. Immunoreactivity to EMA and p53 was focally positive. TTF-1, galectin 3 and thyroglobulin immunoreactivity was restricted to the non-neoplastic thyroid tissue. Both tumor cells and non-neoplastic follicular cells were negative for CD5. The MIB-1 index was 36%. DNA extracted from the tumor identified a BRAF V600E mutation in exon 15 and a BRAF G468A mutation in exon 11, whereas DNA from non-tumorous cells did not contain a mutation. These molecular findings may suggest a direct transformation from papillary carcinoma to SCC-T.