Heying Yang

Discovery and identification of potential biomarkers of papillary thyroid carcinoma.

BackgroundThyroid carcinoma is the most common endocrine malignancy and a common cancer among the malignancies of head and neck. Noninvasive and convenient biomarkers for diagnosis of papillary thyroid carcinoma (PTC) as early as possible remain an urgent need. The aim of this study was to discover and identify potential protein biomarkers for PTC specifically.MethodsTwo hundred and twenty four (224) serum samples with 108 PTC and 116 controls were randomly divided into a training set and a blind testing set. Serum proteomic profiles were analyzed using SELDI-TOF-MS. Candidate biomarkers were purified by HPLC, identified by LC-MS/MS and validated using ProteinChip immunoassays.ResultsA total of 3 peaks (m/z with 9190, 6631 and 8697 Da) were screened out by support vector machine (SVM) to construct the classification model with high discriminatory power in the training set. The sensitivity and specificity of the model were 95.15% and 93.97% respectively in the blind testing set. The candidate biomarker with m/z of 9190 Da was found to be up-regulated in PTC patients, and was identified as haptoglobin alpha-1 chain. Another two candidate biomarkers (6631, 8697 Da) were found down-regulated in PTC and identified as apolipoprotein C-I and apolipoprotein C-III, respectively. In addition, the level of haptoglobin alpha-1 chain (9190 Da) progressively increased with the clinical stage I, II, III and IV, and the expression of apolipoprotein C-I and apolipoprotein C-III (6631, 8697 Da) gradually decreased in higher stages.ConclusionWe have identified a set of biomarkers that could discriminate PTC from non-cancer controls. An efficient strategy, including SELDI-TOF-MS analysis, HPLC purification, MALDI-TOF-MS trace and LC-MS/MS identification, has been proved successful.

Detection and Significance of Serum Protein Marker of Hirschsprung Disease

OBJECTIVE. The objective of this study was to identify a specific fingerprint chromatogram model of serum proteins for early screening and diagnosis of Hirschsprung disease. METHODS. To detect the protein mass spectrograms of 78 serum specimens (42 specimens of Hirschsprung disease, 16 specimens of adhesive ileus including appendicitis and Meckel diverticulum after operation and inflammatory bowel disease, and 20 specimens of normal control subjects), we used surface-enhanced laser desorption/ionization time of flight mass spectrometry technology, combined with bioinformatics methods (support vector machine) to develop and compare protein mass spectrograms from serum samples. RESULTS. We identified 3 protein markers, the mass-to-charge ratio of which is positioned at 3221.7, 5639.2, and 6884.2 from the fingerprint chromatogram model of serum protein for early screening and diagnosis of Hirschsprung disease. The markers had 100% sensitivity and specificity. CONCLUSION. The fingerprint chromatogram model of serum protein using surface-enhanced laser desorption/ionization time of flight mass spectrometry technology combining support vector machine is a new method of early screening and diagnosis of Hirschsprung disease that is worthy of additional research and application.

NDRG1 promotes the multidrug resistance of neuroblastoma cells with upregulated expression of drug resistant proteins.

BACKGROUND Resistance to chemotherapeutic drugs and recurrence are two major causes of poor prognosis in many tumors including neuroblastoma. This study aimed to investigate the effect of the elevated intracellular NDRG1 expression on drug resistance of human neuroblastoma cells to chemotherapy, for exploring novel approaches for biotherapy of neuroblastoma. METHODS Human neuroblastoma KP-N-Ns cell lines were transfected with the lentivirus vector containing human NDRG1 cDNA, with empty vector-transfected or blank cells as controls. Transfection status was confirmed under fluorescence microscope, while PCR assay and western blot analysis were performed to determine the expression changes. MTT and TUNEL assays were used to detect the resistance of the cells to anticancer drugs, including vincristine, cyclophosphamide and so on. Additionally, the expression of drug resistant proteins was detected. RESULTS Stable lentiviral transfection cell line was successfully established with significantly upregulated NDRG1 expression. MTT assay revealed greater cell growth under NDRG1 overexpression with drugs stimulation, as compared to controls. TUNEL assay also showed less apoptosis of NDRG1 overexpressing cells than those of controls when exposed to these drugs, suggesting the increasing drug resistance through NDRG1 overexpression. Besides, the expression of MDR, LRP-1 and MRP-1 was also increased in NDRG1 overexpressing cells, implying NDRG1-mediated pathways in multidrug resistance of neuroblastoma. CONCLUSION NDRG1 could increase the resistance of neuroblastoma cells to chemotherapeutic drugs, with its positive regulation on drug resistant proteins. This study provided new insights for exploring the mechanism of the resistance to chemotherapeutic drugs and also novel approach for biotherapy in neuroblastoma.

Identification of Apolipoprotein C-I as a Potential Wilms’ Tumor Marker after Excluding Inflammatory Factors

Wilms’ tumor is one of the most common malignant tumors observed in children, and its early diagnosis is important for late-stage treatment and prognosis. We previously screened and identified protein markers for Wilms’ tumor; however, these markers lacked specificity, and some were associated with inflammation. In the current study, serum samples from children with Wilms’ tumors were compared with those of healthy controls and patients with systemic inflammatory response syndrome (SIRS). After exclusion of factors associated with inflammation, specific protein markers for Wilms’ tumors were identified. After comparing the protein peak values obtained from all three groups, a protein with a m/z of 6438 Da was specified. Purification and identification of the target protein using high-pressure liquid chromatography (HPLC) and two-dimensional liquid chromatography-linearion trap mass spectrometry(2D-LC-LTQ-MS) mass spectrometry, respectively, revealed that it was apolipoprotein C-I (APO C-I). Thus, APO C-I is a specific protein marker for Wilms’ tumor.

Diagnostic and prognostic role of serum protein peak at 6449 m/z in gastric adenocarcinoma based on mass spectrometry

Background:Gastric cancer (GC) is a highly aggressive cancer type associated with significant mortality owing to delayed diagnosis and non-specific symptoms observed in the early stages. Therefore, identification of novel specific GC serum biomarkers for screening purposes is an urgent clinical requirement.Methods:This study recruited a total of 432 serum samples from 296 GC patients split into the mining and testing sets. We aimed to screen for reliable protein biomarkers from matched serum samples based on mass spectrometry, followed by comparison with three representative conventional markers using receiver operating characteristic and survival curve analyses to ascertain their potential values as diagnostic and prognostic biomarkers for GC.Results:We identified an apoC-III fragment with confirmation in an independent test set from a second hospital. We found that the diagnostic ability of this fragment performed better than current standard GC diagnostic biomarkers both individually and in combination in distinguishing patients with GC from healthy individuals. Moreover, we found that this apoC-III protein fragment represents a more robust potential prognostic factor for GC than the three conventional markers.Conclusions:In view of these findings, we suggest that apoC-III protein fragment is a novel diagnostic and prognostic biomarker, a complement to conventional biomarkers in detecting GC.

Evaluation of promoter hypomethylation and expression of p73 as a diagnostic and prognostic biomarker in Wilms’ tumour

Aims A member of the p53 family, the p73 gene is essential for the maintenance of genomic stability, DNA repair and apoptosis regulation. This study was designed to evaluate the utility of expression and DNA methylation patterns of the p73 gene in the early diagnosis and prognosis of Wilms’ tumour (WT). Methods Methylation-specific PCR, semi-quantitative (sq-PCR), real-time quantitative PCR (qRT–PCR), receiver operating characteristic (ROC), and survival and hazard function curve analyses were utilised to measure the expression and DNA methylation patterns of p73 in WT tissue samples with a view to assessing diagnostic and prognostic value. Results The relative expression of p73 mRNA was higher, while the promoter methylation level was lower in the WT than the control group (p<0.05) and closely associated with poor survival prognosis in children with WT (p<0.05). Increased expression and decreased methylation of p73 were correlated with increasing tumour size, clinical stage and unfavourable histological differentiation (p<0.05). ROC curve analysis showed areas under the curve of 0.544 for methylation and 0.939 for expression in WT venous blood, indicating the higher diagnostic yield of preoperative p73 expression. Conclusions Preoperative venous blood p73 level serves as an underlying biomarker for the early diagnosis of WT. p73 overexpression and concomitantly decreased promoter methylation are significantly associated with poor survival in children with WT.

Diagnostic and prognostic significance of serum apolipoprotein C-I in triple-negative breast cancer based on mass spectrometry.

ABSTRACT Women with triple-negative breast cancer (TNBC) have poor prognosis because of the aggressive nature of the tumor, delayed diagnosis and non-specific symptoms in the early stages. Identification of novel specific TNBC serum biomarkers for screening and therapeutic purposes therefore remains an urgent clinical requirement.We obtained serum samples from a total of 380 recruited individuals split into mining and testing sets, with the aim of screening for reliable protein biomarkers from TNBC and non-TNBC (NTNBC) sera. Samples were assessed using mass spectrometry, followed by receiver operating characteristic (ROC), survival and hazard function curve as well as multivariate Cox regression analyses to ascertain the potential of the protein constituents as diagnostic and prognostic biomarkers for TNBC.We identified upregulated apolipoprotein C-I (apoC-I) with a validated positive effect on TNBC tumorigenesis, with confirmation in an independent test set and minimization of systematic bias by pre-analytical parameters. The apoC-I protein had superior diagnostic ability in distinguishing between TNBC and NTNBC cases. Moreover, the protein presented a more robust potential prognostic factor for TNBC than NTNBC. The apoC-I protein identified in this study presents an effective novel diagnostic and prognostic biomarker for TNBC, indicating that measurement of the peak intensity at 7785 Da in serum samples could facilitate improved early detection and estimation of postoperative survival prognosis for TNBC.

MicroRNA-211 causes ganglion cell dysplasia in congenital intestinal atresia via down-regulation of glial-derived neurotrophic factor

MicroRNAs (miRNAs) are known to be involved in normal brain functions and nervous system diseases. Some evidence have pointed to the dysregulation of miRNAs in congenital intestinal atresia. In this study, we investigated the differential expression of miRNAs and the posttranscriptional regulation of glial‐derived neurotrophic factor (GDNF) by endogenous miRNA in congenital intestinal atresia.

Assessment of promoter methylation and expression of SIX2 as a diagnostic and prognostic biomarker in Wilms’ tumor

This study was designed to evaluate the utility of expression and DNA methylation patterns of the sine oculis homeobox homolog 2 (SIX2) gene in early diagnosis and prognosis of Wilms’ tumor (WT). Methylation-specific polymerase chain reaction (MSP), real-time quantitative polymerase chain reaction (qRT-PCR), receiver operating characteristic (ROC), and survival curve analyses were utilized to measure the expression and DNA methylation patterns of SIX2 in a cohort of WT tissues, with a view to assessing their diagnostic and prognostic value. Relative expression of SIX2 mRNA was higher, while the promoter methylation level was lower in the WT than control group (P < 0.05) and closely associated with poor survival prognosis of WT children (P < 0.05). Increased expression and decreased methylation of SIX2 were correlated with increasing tumor size, clinical stage, vascular invasion, and unfavorable histological differentiation (P < 0.05). ROC curve analysis showed areas under the curve (AUCs) of 0.579 for methylation and 0.917 for expression in WT venous blood, indicating higher diagnostic yield of preoperative SIX2 expression. The preoperative venous blood SIX2 expression level serves as an underlying biomarker for early diagnosis of WT. SIX2 overexpression and concomitantly decreased promoter methylation are significantly associated with poor survival of WT children.

TNF-α induces autophagy through ERK1/2 pathway to regulate apoptosis in neonatal necrotizing enterocolitis model cells IEC-6

ABSTRACT Necrotizing enterocolitis (NEC) is a potentially fatal illness in premature neonates. Tumor necrosis factor-α (TNF-α) and autophagy are associated with the pathogenesis of NEC. This study aimed to explore whether TNF-α might regulate apoptosis in neonatal NEC model cells IEC-6 via regulation of autophagy. NEC rat model was induced by hand feeding and exposure to asphyxia/cold-stress for histologic examination. The NEC in vitro model (IEC-6/NEC cells) was established by stimulating the intestinal epithelial cell line IEC-6 with lipopolysaccharide (LPS, 100 μg/mL) for 3 h to investigate the effects of TNF-α on IEC-6 proliferation and apoptosis. In this study, NEC rats showed decreased proliferating cell nuclear antigen (PCNA) expression, increased TUNEL-positive cells, higher expression of TNF-α, p-ERK1/2, and autophagy-related proteins in rat small intestine compared with their controls. Additionally, the LPS-stimulated IEC-6/NEC cells showed a significantly decreased proliferation and increased apoptosis compared with the control cells. Furthermore, the LPS-stimulated IEC-6/NEC cells exhibited enhanced autophagy level, as evidenced by a dose-dependent increase in Beclin-1 protein expression, LC3II/LC3I ratio and accumulation of MDC-positive autophagic vacuoles. Moreover, inhibition of autophagy by wortmannin or LY294002 significantly abolished the LPS-mediated decreased proliferation and increased apoptosis of IEC-6/NEC cells. Results also showed that inhibition of ERK1/2 pathway using U0126 significantly inhibited TNF-α-induced autophagy. Furthermore, the TNF-α-mediated inhibition of IEC-6 proliferation and promotion of IEC-6 apoptosis was abolished by U0126. Our findings demonstrated that TNF-α might induce autophagy through ERK1/2 pathway to regulate apoptosis in neonatal NEC cells IEC-6. Our study enhances our understanding of neonatal NEC pathogenesis.