scholarly journals Identifying Circulating MicroRNA in Kawasaki Disease by Next-Generation Sequencing Approach

2021 ◽  
Vol 43 (2) ◽  
pp. 485-500
Author(s):  
Ken-Pen Weng ◽  
Ching-Feng Cheng ◽  
Kuang-Jen Chien ◽  
Luo-Ping Ger ◽  
Shih-Hui Huang ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Kawasaki Disease ◽  
Viral Infection ◽  
Intravenous Immunoglobulin ◽  
Healthy Controls ◽  
Next Generation ◽  
Characteristic Analysis ◽  
Expression Levels ◽  
Potential Biomarker ◽  
Generation Sequencing

Kawasaki disease (KD) typically occurs in children aged under 5 years and can cause coronary artery lesions (CALs). Early diagnosis and treatment with intravenous immunoglobulin can reduce the occurrence of CALs; therefore, identifying a good biomarker for diagnosing KD is essential. Here, using next-generation sequencing in patients with recurrent KD, those with viral infection, and healthy controls, we identified dysregulated circulating microRNAs as diagnostic biomarkers for KD. Pathway enrichment analysis illustrated the putative role of these miRNAs in KD progression. Their expression levels were validated using real-time polymerase chain reaction (qPCR). Fifteen dysregulated circulating miRNAs (fold changes >2 and <0.5) were differentially expressed in the recurrent KD group compared with the viral infection and control groups. These miRNAs were significantly involved in the transforming growth factor-β, epithelial–mesenchymal transition, and cell apoptosis signaling pathways. Notably, their expression levels were frequently restored after intravenous immunoglobulin treatment. Among the candidates, miR-24-3p expression level was significantly higher in patients with recurrent KD compared with healthy controls or viral infection controls (p < 0.001). Receiver operating characteristic analysis revealed that high miR-24-3p expression levels may be a potential biomarker for KD diagnosis. In conclusion, we identified miR-24-3p significantly higher in KD patients, which may be a potential diagnostic biomarker for KD.

scholarly journals Prediction for Intravenous Immunoglobulin Resistance Combining Genetic Risk Loci Identified From Next Generation Sequencing and Laboratory Data in Kawasaki Disease

2020 ◽  
Vol 8 ◽  
Author(s):  
Liqin Chen ◽  
Sirui Song ◽  
Qianqian Ning ◽  
Danying Zhu ◽  
Jia Jia ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Kawasaki Disease ◽  
Intravenous Immunoglobulin ◽  
Laboratory Data ◽  
Healthy Children ◽  
Next Generation ◽  
Sequencing Technology ◽  
Scoring Model ◽  
Ivig Resistance ◽  
Generation Sequencing

Background: Kawasaki disease (KD) is the most common cause of acquired heart disease. A proportion of patients were resistant to intravenous immunoglobulin (IVIG), the primary treatment of KD, and the mechanism of IVIG resistance remains unclear. The accuracy of current models predictive of IVIG resistance is insufficient and doesn't meet the clinical expectations.Objectives: To develop a scoring model predicting IVIG resistance of patients with KD.Methods: We recruited 330 KD patients (50 IVIG non-responders, 280 IVIG responders) and 105 healthy children to explore the susceptibility loci of IVIG resistance in Kawasaki disease. A next generation sequencing technology that focused on 4 immune-related pathways and 472 single nucleotide polymorphisms (SNPs) was performed. An R package SNPassoc was used to identify the risk loci, and student's t-test was used to identify risk factors associated with IVIG resistance. A random forest-based scoring model of IVIG resistance was built based on the identified specific SNP loci with the laboratory data.Results: A total of 544 significant risk loci were found associated with IVIG resistance, including 27 previous published SNPs. Laboratory test variables, including erythrocyte sedimentation rate (ESR), platelet (PLT), and C reactive protein, were found significantly different between IVIG responders and non-responders. A scoring model was built using the top 9 SNPs and clinical features achieving an area under the ROC curve of 0.974.Conclusions: It is the first study that focused on immune system in KD using high-throughput sequencing technology. Our findings provided a prediction of the IVIG resistance by integrating the genotype and clinical variables. It also suggested a new perspective on the pathogenesis of IVIG resistance.

Identification and Characterization of a Novel Parvovirus-Like Virus in Seronegative Hepatitis Patients by Next Generation Sequencing

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 273-273
Author(s):  
Baoyan Xu ◽  
Ning Zhi ◽  
Gangqing Hu ◽  
Zhihong Wan ◽  
Sachiko Kajigaya ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Capsid Protein ◽  
Conflicts Of Interest ◽  
Virus Titer ◽  
The United States ◽  
Western China ◽  
Healthy Controls ◽  
Next Generation ◽  
Specific Antibodies ◽  
Generation Sequencing

Abstract Abstract 273 Seronegative hepatitis—non-hepatitis A, non-B, non-C, non-E—is poorly characterized but strongly associated with serious complications, especially aplastic anemia and fulminant hepatitis of childhood. Seronegative hepatitis is rare in the United States but more prevalent in Asia, constituting about 10–20% of acute cases. We applied next-generation sequencing to blood samples of patients from western China with seronegative hepatitis for virus discovery. A total of 92 plasma specimens were collected at Chongqing, China, between 1999 and 2007. Twenty-seven patients were diagnosed as having acute hepatitis by clinical and laboratory characteristics. Sixty-five patients had biopsy-proven chronic aggressive hepatitis, ten of which had cirrhosis. Serologic assays for hepatitis viruses A, B, C, E, HIV, Epstein-Barr virus and cytomegalovirus were all negative. Additional tests for antinuclear antibody, rheumatoid factor, anti-mitochondrial antibody also were normal. Ten plasma pools derived from 93 specimens of the patients were screened by Solexa deep sequencing. We discovered a 3780-bp contig present in all ten-pools that yielded tBLASTx E scores of 0.003 to 1.5 against parvoviruses. The sequence of the in silico assembled 3780-bp contig was confirmed by overlapping PCRs, indicating the contig that contained the nearly complete new virus genome indeed existed in the patient samples rather than being artificially generated by misassembly. The new virus is provisionally designated NIH-CQV. Further analysis revealed that the contig was composed of two major open reading frames (ORF). Protein Blast showed that ORF1 encoded a protein that contained a conserved P-loop NTPase domain, homologous to the replication-associated protein of bat circovirus (E score=4e-04). ORF2 was homologous to capsid protein of porcine parvovirus (E scores=7e-06). Phylogenetic analysis indicated that the NIH-CQV represents a new subfamily of parvovirus, located at the interface of Parvoviridae and Circoviridae (Figure 1). Prevalence of the NIH-CQV in hepatitis patients was investigated by qPCR. Sixty three out of 92 (69%) patient samples were positive, while all 45 healthy controls were negative. The average virus titer in the patients was 1.28 E4 copies/ul, and the highest one was 3.2 E4 copies/ul. Specific antibodies against NIH-CQV were sought by immunoblot using a recombinant capsid protein. No cross reactivity was detected between the capsid protein of NIH-CQV and other major human parvoviruses. Eighty five percent (78/92) of patients were positive for IgG, and 32% (29/92) of them were positive for IgM. In contrast, 78% (35/45) of healthy controls were positive for IgG and 16% (7/45) were positive for IgM. Viral particles were purified from IgM-positive patient plasma by ultracentrifugation through a 40% sucrose cushion and examined by electron microscopy: spherical, naked, parvovirus-like particles approximately 26–29 nm in diameter were visualized. There was no correlation between clinical diagnosis and the presence or absence of the viral DNA or specific antibodies. Although more work is needed to determine the etiologic role of NIH-CQV in human disease, our data indicate that a novel parvovirus-like virus is highly prevalent in a cohort of patients with seronegative hepatitis. Figure 1, whole-proteome tree of the new parvovirus and members of the families Parvoviridae and Circoviridae. Figure 1,. whole-proteome tree of the new parvovirus and members of the families Parvoviridae and Circoviridae. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Lidocaine and Bupivacaine Downregulate MYB and DANCR lncRNA by Upregulating miR-187-5p in MCF-7 Cells

2022 ◽  
Vol 8 ◽  
Author(s):  
Chiao-Yi Lin ◽  
Wen-Ting Tseng ◽  
Yao-Yin Chang ◽  
Mong-Hsun Tsai ◽  
Eric Y. Chuang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Next Generation Sequencing ◽  
Expression Profiles ◽  
Luciferase Reporter ◽  
Functional Assay ◽  
Next Generation ◽  
Potential Biomarker ◽  
Reporter Assays ◽  
Generation Sequencing ◽  
Mcf 7

Background: Breast cancer is the most common malignancy and a leading cause of death among women. The majority of patients require surgery, and retrospective studies have revealed an association between anaesthetic techniques during surgery and clinical outcomes. Local anaesthetics (LAs) influence carcinogenesis by interacting with non-coding RNAs (ncRNAs). However, the detailed mechanisms underlying the association between LAs and ncRNAs remain unclear.Methods: In this study, the effects of two commonly used LAs, lidocaine and bupivacaine, on the malignancy of MCF-7 breast cancer cells were investigated. The expression profiles of the microRNAs (miRNAs) that responded to treatment with LAs were determined through next-generation sequencing.Results: Data from the functional assay revealed that the LAs suppressed the proliferation of MCF-7 cells. The result of next-generation sequencing revealed that 131 miRNAs were upregulated, following treatment with the LAs. Validation using polymerase chain reaction (PCR) identified miR-187-5p as a potential biomarker, and it was selected for further analyses. Prediction with bioinformatics tools and luciferase reporter assays revealed that MYB is a direct target gene of miR-187-5p. Based on the hypothesis that lncRNAs acts as miRNA sponges, the target lncRNA, DANCR, of miR-187-5p was predicted using DIANA-LncBase v2 and validated using luciferase reporter assays. In addition, the reciprocal suppressive effect between DANCR and miR-187-5p was determined.Conclusions: This study suggests that one of the anti-tumour mechanisms of lidocaine and bupivacaine is mediated through the DANCR-miR-187-5p-MYB axis. This may provide a novel molecular mechanism of tumour suppression in breast cancer.

scholarly journals Genotypic Characteristics of Hepatoblastoma as Detected by Next Generation Sequencing and Their Correlation With Clinical Efficacy

2021 ◽  
Vol 11 ◽  
Author(s):  
Huimin Hu ◽  
Weiling Zhang ◽  
Tian Zhi ◽  
Jing Li ◽  
Yuan Wen ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Survival Rate ◽  
Mismatch Repair ◽  
Pediatric Patients ◽  
Next Generation ◽  
Expression Levels ◽  
Dna Mismatch ◽  
Mismatch Repair Status ◽  
Generation Sequencing ◽  
Potential Biomarkers

BackgroundHepatoblastoma (HB) is the most common malignant embryonic liver tumor type in children under 3 years of age. In the present study, the next generation sequencing (NGS) method was used to detect the genotype characteristics of HB and summarize the correlation between the common mutation genotypes noted in this disease and the clinical treatment and prognosis. The results may aid clinical prognosis and the successful application of targeted drugs.MethodsInitially, DNA was extracted from tumor tissue specimens and peripheral blood derived from 19 pediatric patients with HB. Subsequently, DNA panel and NGS methods were used to detect tumor diagnosis and the expression levels of treatment-associated genes, followed by the summary of genotype characteristics. In addition, in order to further assess the application of immunotherapy in HB, immunohistochemical detection of programmed cell death 1 ligand 1 (PDL1) was performed in combination with tumor mutation burden (TMB) and DNA mismatch repair status analysis. Furthermore, the clinical treatment effect and prognosis of the pediatric patients were statistically analyzed according to the characteristics of the genotype. Overall prognosis and prognostic analyses in different groups were performed by Kaplan-Meier and log-rank tests, respectively. Finally, expression validation and diagnostic analysis of commonly reported genes were performed in the GSE75271 dataset, which was obtained from the Gene Expression Omnibus (GEO) database.ResultsIn the present study, certain mutated genes, including nuclear factor erythroid 2-related factor 2 (NFE2L2), catenin β1 (CTNNB1), MYCN, tumor protein p53, axis inhibition protein 1 (AXIN1) and adenomatous polyposis coli (APC) were associated with the pathogenesis of HB. During TMB and DNA mismatch repair status analyses, pediatric patients had a low TMB. All of them did not present with microsatellite instability. The immunohistochemical results indicated lower expression levels of PDL1 in HB. The complete remission (CR) rate of pediatric patients in the gene abnormality group was lower than that of the non-reported disease-associated gene abnormality group. The 2-year overall survival rate and disease-free survival rate of 19 pediatric patients with HB were 72.1% and 42.4%, respectively. Receiver operating characteristic (ROC) analysis demonstrated that CTNNB1, NFE2L2, AXIN1, APC, MYCN and insulin growth factor 2 (IGF2) may be potential biomarkers that could be used for the diagnosis of HB.ConclusionThe genotype changes in HB were more common and the CR rate of the pediatric patients with an altered genotype was lower than that of pediatric patients without an altered genotype. In addition, pediatric patients with HB exhibited lower TMB compared with adult patients. Moreover, the data indicated that CTNNB1, NFE2L2, AXIN1, APC, MYCN and IGF2 may be potential biomarkers that can be used for the diagnosis of HB.

FGFR expression, fusion and mutation as detected by NGS sequencing of DNA and RNA.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e16061-e16061
Author(s):  
Ivan De Dios ◽  
Wanlong Ma ◽  
Spiraggelos Antzoulatos ◽  
Jeffrey Estella ◽  
Maher Albitar
Keyword(s):  
Lung Cancer ◽  
Next Generation Sequencing ◽  
Next Generation ◽  
Lung Cancers ◽  
Expression Levels ◽  
Dna And Rna ◽  
Significant Difference ◽  
High Incidence ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

e16061 Background: Fibroblast Growth Factor Receptors (FGFR1-4) abnormalities (fusion, amplification and mutations) are common in urothelial, breast and endometrial cancers. However, FGFR1-4 have been shown to play a major role in cell proliferation, differentiation, and apoptosis in other types of cancers including colorectal (CRC) and lung cancers. We explored the value of using DNA and RNA next generation sequencing (NGS) in determining the presence of abnormalities in FGFR1-4 in various types of cancer. Methods: Using targeted panel and next generation sequencing (NGS), we analyzed DNA sequencing data (434 genes) in 438 Solid tumors and RNA data (1408 genes) in 160 lung cancers and 53 colorectal cancers (CRC). The expression levels of the CRC and lung cancer were also compared with expression levels of 32 cases of endometrial, urothelial and breast cancers as a group of cancers known to have high incidence. Results: The DNA data showed mutations in 85 samples and CNV in 12 samples. The detected mutations were 18% in FGFR1, 25% in FGFR2, 45% in FGFR3, and 12% in FGFR4. Only 20% of the detected mutations by NGS testing can be detected if the PCR-based FDA-approved kit was used. Analysis of the expression levels of FGFR1-4 mRNA in CRC and lung cancer showed highest expression in FGFR2, followed by FGFR1 then FGFR3. Expression of FGFR4 was the lowest (P < 0.0001). There was no difference between CRC and lung cancer in FGFR1 and FGFR2 mRNA, but FGFR3 was slightly higher in lung cancer as compared with CRC (P = 0.01). FGFR4 was significantly higher in CRC as compared with lung cancer (P < 0.0001). No fusion involving FGFR1-4 was detected in any of the tested CRC or lung cancers. Upon comparing overall expression between CRC/lung cancer with the group of cancers that are known to have high incidence of FGFR1-4 abnormalities (urothelial, breast, and endometrial), FGFR1 and FGFR2 mRNA were significantly lower in CRC/lung cancers (P < 0.0001 and P = 0.0002, respectively), but there was no significant difference in FGFR3. However, significant overlap is noted. In contrast, FGFR4 was significantly higher in CRC (P < 0.0001). Conclusions: This data suggests that while FGFR1-3 genes are overall expressed in CRC and lung, some cases may have significantly high expression of FGFR1-3 and perhaps these cases should be singled out for treatment with FGFR inhibitors. Furthermore, NGS testing for mutations significantly more efficient and can detect significant number of mutations that can be missed if PCR-based testing is used. NGS testing of DNA and RNA is the most appropriate testing for abnormalities in FGFR1-4.

scholarly journals Next-Generation Sequencing Reveals Downregulation of the Wnt Signaling Pathway in Human Dysmature Cumulus Cells as a Hallmark for Evaluating Oocyte Quality

2020 ◽  
Vol 1 (3) ◽  
pp. 205-215
Author(s):  
Ryosuke Akino ◽  
Daisuke Matsui ◽  
Ryouka Kawahara-Miki ◽  
Mitsuyoshi Amita ◽  
Kuniko Tatsumi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Next Generation Sequencing ◽  
Expression Patterns ◽  
Cumulus Cell ◽  
Cumulus Cells ◽  
Oocyte Quality ◽  
Next Generation ◽  
Expression Levels ◽  
Morphological Evaluation ◽  
Generation Sequencing

Background: Dysmature cumulus cells are lower fertilization rates and abnormalities in embryonic development compared to maturation cumulus cells. Morphological evaluation of cumulus–oocyte complexes (COCs) considered the possibility that differences may also be found in gene expression. Purpose: To identify hallmarks for evaluating oocyte quality by investigating gene expression patterns in human cumulus cells surrounding oocytes. Methods: Cumulus cells were obtained from the cumulus–oocyte complex of infertile women treated with assisted reproductive technology. Based on maturity level, the cumulus cells were classified into two categories, i.e., dysmature cumulus cell (DCC) and maturation cumulus cell. DCCs were subjected to gene expression analysis using next-generation sequencing and compared with COCs that are in the process of maturation as controls. Results: The expression levels of genes involved in the Wnt signal/β-catenin pathway were significantly reduced in DCCs compared with those in control cells. Moreover, the expression levels of genes involved in multiple pathways associated with apoptosis were also significantly reduced compared with those in control cells. Conclusions: DCCs showed significant decreases in apoptosis- and Wnt/β-catenin signaling-associated gene expression. DCCs could be classified by morphological evaluation, and the method described in this study may be useful as an oocyte quality estimation tool.

scholarly journals Prevalence and coexistence of KRAS, BRAF, PIK3CA, NRAS, TP53, and APC mutations in Indian colorectal cancer patients: Next-generation sequencing–based cohort study

Tumor Biology ◽  
2017 ◽  
Vol 39 (2) ◽  
pp. 101042831769226 ◽  
Author(s):  
Mayank Jauhri ◽  
Akanksha Bhatnagar ◽  
Satish Gupta ◽  
Manasa BP ◽  
Sachin Minhas ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Next Generation Sequencing ◽  
Cancer Patients ◽  
Gene Mutations ◽  
Next Generation ◽  
Kras Mutations ◽  
Pik3ca Mutations ◽  
Potential Biomarker ◽  
Colorectal Cancer Patients ◽  
Generation Sequencing

Colorectal cancer incidences are on a rise in India. In this study, we have analyzed the mutation frequencies of six potential biomarkers, their coexistence, association with clinicopathological characteristics, and tumor location in Indian colorectal cancer patients. Next-generation sequencing was performed to identify mutations in the six potential biomarker genes using formalin-fixed paraffin-embedded tissue blocks of 112 colorectal cancer patients. The mutation frequency observed in KRAS, BRAF, PIK3CA, NRAS, TP53, and APC was 35.7%, 7.1%, 16.1%, 6.3%, 39.3%, and 29.5%, respectively. The significant associations of mutations were KRAS with age less than 60 years (p = 0.041), PIK3CA with males (p = 0.032), tumor stage I–II (p = 0.013), lack of metastasis in lymph nodes (p = 0.040), NRAS with rectum (p = 0.002), and APC with T2 stage of tumor growth (p = 0.013). No single patient harbored mutations in these six genes or any five genes simultaneously. Significance was noted in coexistence of KRAS with APC (p = 0.024) and mutual exclusion of KRAS with BRAF (p = 0.029). PIK3CA exon 9 was observed to be more frequently associated with KRAS mutations than PIK3CA exon 20 (p = 0.072). NRAS mutations were mutually exclusive with BRAF and PIK3CA mutations. As per our knowledge, this is the first next-generation sequencing–based biomarker study in Indian colorectal cancer patients. Frequent coexistence of gene mutations in pairs and triplets suggests that synergistic effect of overlapping mutations might further trigger the disease. In addition, infrequent coexistence of multiple gene mutations hints toward different signaling pathways for colorectal cancer tumorigenesis.

scholarly journals Next-generation sequencing identifies micro-RNA–based biomarker panel for Kawasaki disease

2016 ◽  
Vol 138 (4) ◽  
pp. 1227-1230 ◽  
Author(s):  
Ho-Chang Kuo ◽  
Kai-Sheng Hsieh ◽  
Mindy Ming-Huey Guo ◽  
Ken-Pen Weng ◽  
Luo-Ping Ger ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Kawasaki Disease ◽  
Micro Rna ◽  
Next Generation ◽  
Biomarker Panel ◽  
Generation Sequencing
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