Objective To analyze the metabolic profiles of the female papillary thyroid carcinoma (PTC) and the relationship between the metabolic profiles and primary tumor size and cervical lymph node metastasis using a metabolomics approach.Methods Forty-three cases of female PTC were enrolled in this study.Gas chromatography-time-of-flight mass spectrometry and ultra performance liquid chromatography-time of flight-mass spectrometry were employed for analyzing metabolic profiles of tumor tissues and adjacent normal tissues in the female PTC.Cases were divided into Group T1 (tumor size ≤2.0 cm) and Group T2 (tumor size ＞ 2.0 cm) according to the tumor size,and divided into Group N-(with negative cervical lymph node) and Group N + (with positive cervical lymph node) according to the cervical lymph node conditions.We compared the metabolomic profiles between these groups.Results A panel of 46 differentially expressed metabolites was identified in the PTC specimens,compared with normal tissues.Increased metabolism of amino acid,purine and pyrimidine,tryptophan acid,one carbon,glycolysis,taurine and hypotaurine,and fatty acid were found in PTC tumors tissues.Amino acids,purine and pyridine,tryptophan,and carbon metabolism increased significantly in the tumor tissues of Group T2 compared with Group T1,while glycolysis,amino acid,purine and pyridine,tryptophan,and carbon metabolism increased significantly in the specimens of Group N +.Conclusion Distinct metabolic profiles were identified in the female PTC tissues,which were related to the primary tumor size and cervical lymph node metastases.

Objective:To identify important prognostic molecular markers of triple negative breast cancer (TNBC) using high throughput sequencing technology and to explore the correlation of spindle checkpoint protein BUB1B and clinicopathological features with patients′ prognosis.Methods:The clinicopathological data and prognostic information of TNBC diagnosed at the West China Hospital of Sichuan University from 2009 to 2017 were collected. Forty-seven fresh tumor samples and 139 formalin fixed paraffin-embedded samples were selected. The fresh tumor samples were subject to RNA sequencing (RNA-seq). The enrichment analysis and protein-protein interaction (PPI) analysis were performed after intersection of difference analysis between RNAseq and GEO (Gene Expression Omnibus) datasets GSE38959 and GSE65194. Kaplan-Meier plotter database was used to analyze the relationship between expression of BUB1B and prognosis. Immunohistochemical staining was used to verify its expression in TNBC and correlation with clinicopathological features and prognosis.Results:Using edgeR to perform differential expression analysis between 47 TNBC tumor tissues and 12 normal tissues, 1 559 up-regulated genes and 1 376 down-regulated genes were identified, while only 131 differentially expressed genes were overlapping with those in GSE38959 and GSE65194. Enrichment analysis was mainly enriched in cell cycle, JAK-STAT signaling pathway and p53 signaling pathway. The top 10 genes ranked by degree of association were TOP2A, BUB1B, MKI67, PLK1, RRM2, PCNA, KPNA2, SMC4, PBK and IGF1. Kaplan-Meier plotter database analysis showed that the expression of BUB1B was significantly correlated with the prognosis of TNBC [overall survival, hazard ratio (HR)=0.52, 95% CI (0.35-0.77), P=0.001; distant metastasis-free, HR=0.72, 95% CI (0.52-0.98), P=0.038]. The immunohistochemical analyses of 139 formalin fixed paraffin-embedded samples showed that the low expression of BUB1B was correlated with poor prognosis in TNBC [HR=0.41, 95% CI (0.18-0.95), P=0.024]. Conclusions:The low expression of BUB1B protein is associated with poor prognosis in TNBC patients, and the molecular mechanism related with prognosis and potential therapeutic targets need to be further studied.

To obtain chicken CD40L protein, the cDNA was prepared from chicken splenic cells and used as a template to clone and amplify CD40L by PCR. The target gene was cloned into pFastBac vector to construct a pFastBac-chCD40L donor plasmid. Recombinant plasmid was transformed into DH10Bac and recombinant Bacmid-chCD40L was obtained. The Bacmid-chCD40L plasmid was transfected into sf9 insect cells to obtain His-chCD40L protein. In addition, the target gene was cloned into pQM01 vector to construct a pQM01-chCD40L plasmid, recombinant plasmid was transfected into HEK 293T cells to obtain Strep-chCD40L protein. The chCD40L protein was purified by affinity chromatography, and the concentration of purified chCD40L protein was determined to be 0.01 mg/mL. Primary cells were isolated from the bursal tissue of 3-week old SPF chickens, and the chCD40L protein was added to the culture medium to stimulate cells. The chCD40L could bind to CD40 on B cells as examined by Western blotting, indirect immunofluorescence assay and flow cytometry, suggesting that chCD40L protein is biologically active. We successfully obtained chicken CD40L protein of biological activity, which laid the foundation in the in vitro culture of primary B lymphocytes for the isolation and diagnosis of virulent IBDV.
Animals ; Baculoviridae/genetics* ; CD40 Ligand/genetics* ; Chickens ; Cloning, Molecular ; Genetic Vectors/genetics* ; Recombinant Proteins/genetics*