Humans ; In Situ Hybridization, Fluorescence ; Tissue Array Analysis ; methods

Objective To investigate clinical characteristics,quality of life,and psychological factors in functional dyspepsia (FD) patients with weight loss.Methods A multi-center and prospective investigation was conducted (Clinical trial registration number:ChiCTR-TRC-12001968) which recruited 1341 consecutive FD outpatients from three Guangdong hospitals according to Rome Ⅲ criteria between June and September in 2012.Of these,1057 patients with complete information were divided into group A as having weight loss more than 5％ (n =207) and group B as having weight loss less than 5％ (n =850).Nepean Dyspepsia Index-symptom checklist,appetite,Hamilton Rating Scale of Anxiety/Depression (HAMA/HAMD),Pittsburgh Sleep Quality Index (PSQI),Nepean Dyspepsia Index-quality of life,direct medical costs were used to compare the differences between the two groups.Results There were no significant differences in total scores of NDI-symptom checklist,the severity or bothersome items between two groups (t =-1.301、-0.918、0.138,P =0.193、0.359、0.890),but significant differences were observed in the frequence item and the rating of appetite as poor or very poor between two groups (t =-2.122,P =0.035 ;x2 =35.448,P =0.000 ;x2 =35.274,P =0.000).Significant differences were observed in the incidence of anxiety and depression and total scores of PSQI between the two groups (x2 =73.939,P =0.000;x2 =47.046,P =0.000 ; t =-4.904,P =0.000).Subscales scores of NDI-quality of life in group B were all significantly higher than those in group A (t =5.348、2.569、5.809、4.704,P =0.000、0.010、0.000、0.000).There were significant differences in the frequency of care-seeking and direct medical costs between the two groups (t =-4.860、-3.011,P =0.000、0.003).The frequency of gastroscopic examinations in group A was slightly higher than that in group B (t =-1.505,P =0.133).Conclusion FD patients with weight loss is not rare,and most of them have psychological disorders,loss of appetite and sleep disturbance.Moreover,care-seeking are more frequent,direct medical costs are more expensive,and impairment as assessed by NDI-QOL is more severe.Therefore,we should pay more attention to FD with weight loss in clinical setting.

Objective: To investigate epidermal growth factor receptor (EGFR) mutation status in lung adenocarcinomas before and after acquiring resistance to EGFR tyrosine kinase inhibitors (TKIs) using ARMS method followed by further verification using droplet digital PCR technique. Methods: Twenty qualified patients were included, among them 13 were male and 7 were female patients. Before EGFR-TKIs treatment, 5 patients were EGFR wild-type by ARMS, and the other 15 patients had L858R or 19-del point mutations. The time to progression varied from 4 to 18 months. Mutation of exons 18, 19, 20 and 21 were detected by ARMS, and were verified by droplet digital PCR system method. Results: EGFR wild-type status was unchanged before and after acquired resistance to EGFR-TKIs in 5 lung adenocarcinoma patients. Alteration of EGFR mutation status occurred in 10 of the 15 patients with pre-treatment L858R or 19-del mutations. Among them, T790M mutation was found in 8 patients, L858R became G719X plus S768I mutation in one patient, and 19-del converted into wild-type in one other patient. Conclusions T790M mutation is the primary type of EGFR mutation in lung adenocarcinomas with acquired resistance to EGFR-TKIs therapy. Acquired resistance to EGFR-TKIs dose not lead to the alteration of EGFR status in pre-treatment EGFR wild-type patients, but can alter EGFR mutation status in pre-treatment EGFR mutant patients.

Objective: To compare amplification refractory mutation system(ARMS) and droplet digital PCR (ddPCR) in the detection of epidermal growth factor receptor (EGFR) gene mutations in patients with non-small cell lung cancer (NSCLC), and to investigate the clinical value of ddPCR. Methods: A total of 79 specimens of NSCLC, including 22 cases of cell block, 18 cases of surgical specimens, 12 cases of biopsy specimens and 27 cases of plasma samples, were analyzed for the mutation status of EGFR gene by ARMS and droplet digital PCR method. Results: In 18 cases of surgical specimens and 12 cases of biopsy specimens, the detection results by the two methods were identical with positive rates of 9/18 and 5/12, respectively. In 22 cases of effusion cell blocks, ARMS detected 19-del and L858R of EGFR gene in two cases, in which droplet digital PCR detected 19-del+ T790M mutations in one case and L858R+ T790M mutation in another. L858R mutation was detected by droplet digital PCR in one case but ARMS assay was negative. The remaining 19 cases were consistent by the two methods. In blood samples, the positive rate was 33.3%(9/27) by ARMS and 37.0%(10/27) by droplet digital PCR. Two cases showed L858R and 19-del+ T790M mutation by droplet digital PCR but ARMS assay detected only 19-del. The remaining 25 cases were consistent by the two methods. Conclusion Droplet digital PCR method is more sensitive and accurate than ARMS for the detection of EGFR mutations in pleural fluid and blood samples, can be used in clinical test.

Objective: To study the molecular features of metanephric adenoma (MA) and discuss their values in differential diagnosis. Methods: BRAF V600E immunohistochemistry (IHC) using the mutation-specific VE1 monoclonal antibody and Sanger sequencing of BRAF mutations were performed on 21 MAs, 16 epithelial-predominant Wilms tumors (e-WT) and 20 the solid variant of papillary renal cell carcinomas (s-PRCC) respectively. p16 protein was detected by IHC also. Fluorescence in situ hybridization (FISH) analyses using centromeric probes for chromosome 7 and 17 were performed on the three renal tumors in parallel. Results: Fourteen (14/21, 66.7%) of 21 MA cases demonstrated diffuse, moderate to strong cytoplasmic BRAF V600E IHC staining and the BRAF V600E protein expression was detected in 2 (2/16) of 16 e-WT cases for the first time, whereas all s-PRCCs were negative (P<0.05). All cases (including 14 MAs and 2 e-WTs) with diffuse, moderate to strong cytoplasmic BRAF V600E IHC staining were confirmed to harbor BRAF V600E missense mutations using Sanger sequencing, and no BRAF mutations were detected in cases with negative BRAF V600E protein expression. One case (1/21, 4.8%) showed trisomy of chromosome 7 alone, and another one (1/21, 4.8%) showed trisomy of chromosome 17 alone in 21 MAs. Two cases (2/16) of 16 e-WTs showed trisomy of chromosome 17 alone. In 20 s-PRCCs, trisomy of chromosomes 7 alone was reported in 2 cases (2/20), trisomy of chromosome 17 alone in 3 cases (3/20) and trisomy of chromosome 7 and 17 in 14 cases (14/20). The total positive rates of trisomy of chromosome 7 and/or 17 in MAs, e-WTs and s-PRCCs were 9.6% (2/21), 2/16 and 95.0% (19/20). p16 protein was positive in 81.0% (17/21) MAs, whereas the positive rates in e-WTs and s-PRCCs were 2/16 and 5.0% (1/20). Conclusions Most MAs harbor BRAF V600E mutations, and MAs lack the gains of chromosome 7 and 17 that are characteristic of papillary renal cell carcinoma. These molecular features can be used to distinguish MA from its mimics. BRAF V600E IHC using the mutation-specific VE1 monoclonal antibody provides an effective method in BRAF V600E mutations detection of renal tumors. p16 is overexpressed in MA, and the finding suggests that the low proliferative rate of the tumor might be attributed to BRAF V600E-induced senescence mediated by p16.

Objective: To investigate the clinical, histologic and immunophenotypic features, genetic alterations and prognosis of the rare Xp11 neoplasm with melanocytic differentiation. Methods: Twenty-one cases were selected from the Department of Pathology, Jingling Hospital, Nanjing University School of Medicine from May 2008 to May 2018. The clinicopathologic, immunohistochemical, molecular analysis and follow-up details were collected. Results: There were 7 males and 14 females, with their ages ranging from 4 to 57 years (mean 32.8 years). The tumors were located in kidney (11 cases), pelvis (three cases), and in pancreas, retroperitoneum, adrenal gland, small intestine, prostate, cervix and appendix (one case each). Microscopically, most tumors shared similar morphology such as purely nested or sheet-like architectures separated by a delicate vascular network, purely epithelioid cells with clear to granular eosinophilic cytoplasm, lacks of papillary structures, spindle cell or fat components, uniform round to oval nuclei with small visible nucleoli, and in most of them (16/21) melanin pigment. Immunohistochemically, all cases showed moderately (2+) or strongly (3+) positive staining for TFE3 and Cathepsin K. HMB45 and Melan A were focally expressed in three of 21 cases, while the remaining cases showed typically moderate(2+) or strong (3+) expression. None of the cases were immunoreactive for SMA, desmin, CKpan, S-100 and PAX8. All cases showed TFE3 rearrangement using fluorescence in-situ hybridization (FISH). Fusion FISH assays detected SFPQ-TFE3 gene fusion in 16 cases, NONO-TFE3 gene fusion in two, ASPL-TFE3 and MED15-TFE3 gene fusions in one case each. Polymerase chain reaction and direct sequencing detected SFPQ-TFE3 gene fusion in nine cases, NONO-TFE3 and MED15-TFE3 gene fusions in one case each. Clinical follow-up was available for 15 patients for 12 to 74 months. Six patients died of the disease; and three had recurrences and/or metastases. Six patients were alive with no evidence of disease after initial resection. Conclusions Xp11 neoplasm with melanocytic differentiation has unique morphologic, immunophenotypic and genetic characteristics. The tumor is aggressive, and should be differentiated from Xp11 translocation RCC and perivascular epithelioid cell tumor.