Exploring and comparing of the gene expression and methylation differences between lung adenocarcinoma and squamous cell carcinoma

Lung cancer is one of the most frequently diagnosed malignant tumors and the main reason for cancer-related death around the world, whereas nonsmall cell lung cancer that consists two subtypes: lung adenocarcinoma (LUAD) and squamous cell carcinoma (LUSC) is responsible for an estimated 85% of all lung cancers. The current study aimed to explore gene expression and methylation differences between LUAD and LUSC. EdgeR was used to identify differentially regulated genes between normal and cancer in the LUAD and LUSC extracted from The Cancer Genome Atlas (TCGA), respectively, whereas SAM was used to find genes with differential methylation between normal and cancer in the LUAD and LUSC, respectively. Finally, Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis was performed to analyze the function which these genes enriched in. A total of 391 genes with opposite methylation patterns in LUAD and LUSC and four functional pathways were obtained (false discovery rate (FDR) < 0.1). These pathways mainly included fat digestion and absorption, phenylalanine metabolism, bile secretion, and so on, which were related to the airframe nutrition metabolic pathway. Moreover, two genes CTSE (cathepsin E) and solute carrier family 5 member 7 (SLC5A7) were also found, among which CTSE was overexpressed and hypomethylated in LUAD corresponding to normal lung tissues, whereas SLC5A7 showed the opposite in LUAD. In conclusion, this study investigated the differences between the gene expression and methylation patterns in LUAD and LUSC, and explored their different biological characteristics. Further understanding of these differences may promote the discovery and development of new, accurate strategies for the prevention, diagnosis, and treatment of lung cancer.     

Prognostic value of metabolic genes in lung adenocarcinoma via integrative analyses

BackgroundLung adenocarcinoma (LUAD) is the most common malignant lung tumor. Metabolic pathway reprogramming is an important hallmark of physiologic changes in cancers. However, the mechanisms through which these metabolic genes and pathways function in LUAD as well as their prognostic values have not been fully established.MethodsFour publicly available datasets from GEO and TCGA were used to identify differentially expressed genes (DEGs) in LUAD, which were then subjected to GO and KEGG pathway enrichment analysis. Associations between metabolic gene expressions with overall survival, tumor stage, TP53 mutation status, and infiltrated immune cells were investigated. Protein-protein interactions were evaluated using GeneMANIA and Metascape.ResultsBy integrating four public datasets, 247 DEGs were identified in LUAD. These DEGs were significantly enriched in regulation of chromosome segregation, centromeric region, and histone kinase activity GO terms, as well as in cell cycle, p53 signaling pathway, metabolic pathways, and other KEGG pathways. Elevated expressions of ten metabolic genes in LUAD were significantly associated with poor survival outcomes. These metabolic genes were highly expressed in more advanced tumor stage and TP53 mutated patients. Moreover, expression levels were significantly correlated with tumor-infiltrating immune cells. PPI interaction analysis revealed that the top 20 genes interacting with each metabolic gene were significantly enriched in DNA replication, response to radiation, and central carbon metabolism in cancer.ConclusionThis study elucidates on molecular changes in metabolic genes in LUAD, which may inform the development of genetically oriented diagnostic approaches and effective treatment options.     

家蚕5龄幼虫精巢和卵巢microRNA芯片及转录组比较分析

【目的】本研究旨在通过对家蚕Bombyx mori 5龄幼虫精巢和卵巢组织微小RNA (microRNA, miRNA)基因芯片及转录组进行分析,找到参与家蚕性腺发育相关的miRNA分子及可能的靶基因。【方法】采用新一代高通量测序平台对家蚕5龄幼虫精巢和卵巢(分别定义为Test和Control)进行miRNA基因芯片检测及转录组测序分析,根据P＜0.05且log₂(fold change, FC)≥2的标准,通过比较筛选出Test vs Control的差异表达miRNA;根据q≤0.05且|log₂(fold change)|≥1的标准,通过比较筛选出Test vs Control的差异表达基因 (differentially expressed genes, DEGs);随机选取8个上调和12个下调差异表达miRNA,对其表达及其预测的5个靶基因进行qRT-PCR验证;对DEGs以及差异表达miRNA的靶基因进行KEGG通路富集分析。【结果】从精巢和卵巢样本中(Test vs Control)分别鉴定出68个差异表达miRNA和3 991个DEGs,其中上调和下调miRNA分别为36和32个,上调和下调DEGs分别为2 033和1 958个。差异表达miRNA的qRT PCR验证结果均与芯片数据一致。KEGG通路富集分析结果显示DEGs在新陈代谢及核糖体的信号通路显著富集。对差异表达miRNA在DEGs中的可能靶基因进行预测,结果找到了4组表达趋势相反的miRNA与靶基因：分别是bmo-miR-2774a与LOC101745556;bmo-miR-92b与LOC101735954以及bmo-miR-3266与LOC733130和LOC778467;1组表达趋势一致的miRNA与靶基因：bmo-miR-3321与LOC101744895。5个靶基因的qRT-PCR验证结果与转录组测序结果一致。【结论】本研究获得了家蚕5龄幼虫精巢和卵巢转录组及miRNA芯片数据,筛选并验证了4组差异表达和1组一致表达miRNA及潜在靶基因,为探究家蚕精巢和卵巢发育差异奠定了基础。     

高通量测序技术分析急性髓系白血病血浆外泌体微RNA表达谱差异

急性髓系白血病（acute myeloid leukemia,AML）是一类造血干细胞的恶性克隆性疾病,目前的诊断方法不利于疾病的早期发现,且诊断结果重复性较差。已有大量研究显示,细胞外microRNA（miRNA或miR）富集在外泌体（exosome）中,且受其表面膜的保护而具有很好的稳定性,是理想的分子标志物。目前,多种实体肿瘤均已检测到肿瘤特异性外泌体miRNA(exosomal miRNA)。然而,在AML患者中未见此外泌体miRNA报道。本研究探讨急性髓系白血病血浆外泌体miRNA表达谱差异及新miRNA序列。采用solexa高通量测序技术对7例AML患者（AML组）及7例健康对照者（对照组）血浆外泌体miRNAs进行测序,利用Mireap预测软件进行新miRNAs分析,通过edger差异分析软件筛选组间差异miRNA,获得211个已知的差异表达miRNAs以及2个新miRNAs,选择4个差异表达的miRNAs：miR-155-5p、miR-335-5p、miR-451a及xxx-m0038 5p（新miRNA）,在两组（各23例）的血浆外泌体样本中,进行实时荧光定量PCR（qRT-PCR）验证,验证结果与测序结果一致。对差异表达的外泌体miRNA进行靶基因预测及其GO（Gene Ontology）和信号通路富集分析,发现靶基因聚集的生物学功能多数参与生物进展过程的调控。靶基因主要富集在FoxO、MAPK、Hippo信号通路以及HTLV-I感染等。结果显示,AML患者与健康对照者的血浆外泌体miRNA存在着差异性表达。差异性表达的miRNA特异性很高,对进一步阐明AML白血病发生与发展的分子机制、研发新的无创诊断方法、新的诊断标记物和有效治疗AML的方法具有十分重要和深远的意义。     

Investigation of key microRNAs associated with hepatocellular carcinoma using small RNA-seq data

To identify key microRNAs (miRNAs) associated with hepatocellular carcinoma (HCC) using small RNA-seq data. Small RNA-seq data for two HCC samples and two normal samples were downloaded from NCBI Gene Expression Omnibus. MiRNAs were identified through database search. Differentially expressed miRNAs were screened out with t test and their target genes were retrieved. Functional enrichment analysis was performed to uncover their biological functions. Regulatory networks and core metabolic networks were also constructed to present the global patterns. In addition, new miRNAs and their target genes were predicted. A total of 59 differentially expressed miRNAs were obtained, 12 up-regulated and 47 down-regulated. A total of 3,306 target genes were retrieved for eight miRNAs. Pathway enrichment analysis for the target genes showed that “pathways in cancer” and “MAPK signaling pathway” were significantly over-represented. Functional enrichment analysis found that “biological regulation” and “macromolecule modification” were significantly related to the target genes. Two regulatory networks were constructed for up- and down-regulated differentially expressed miRNAs with information from Ingenuity Pathway Analysis database. Two metabolic networks were also established based upon “pathways in cancer” and “MAPK signaling pathway”, consisting of miRNAs, target genes, compounds and others genes. Moreover, a number of new miRNAs and relevant target genes were predicted. Our study discloses a number of miRNAs as well as genes which may be involved in the development of HCC and these findings are beneficial in guiding future researches.     

Systematic profiling of invasion-related gene signature predicts prognostic features of lung adenocarcinoma

Due to the high heterogeneity of lung adenocarcinoma (LUAD), molecular subtype based on gene expression profiles is of great significance for diagnosis and prognosis prediction in patients with LUAD. Invasion-related genes were obtained from the CancerSEA database, and LUAD expression profiles were downloaded from The Cancer Genome Atlas. The ConsensusClusterPlus was used to obtain molecular subtypes based on invasion-related genes. The limma software package was used to identify differentially expressed genes (DEGs). A multi-gene risk model was constructed by Lasso-Cox analysis. A nomogram was also constructed based on risk scores and meaningful clinical features. 3 subtypes (C1, C2 and C3) based on the expression of 97 invasion-related genes were obtained. C3 had the worst prognosis. A total of 669 DEGs were identified among the subtypes. Pathway enrichment analysis results showed that the DEGs were mainly enriched in the cell cycle, DNA replication, the p53 signalling pathway and other tumour-related pathways. A 5-gene signature (KRT6A, MELTF, IRX5, MS4A1 and CRTAC1) was identified by using Lasso-Cox analysis. The training, validation and external independent cohorts proved that the model was robust and had better prediction ability than other lung cancer models. The gene expression results showed that the expression levels of MS4A1 and KRT6A in tumour tissues were higher than in normal tissues, while CRTAC1 expression in tumour tissues was lower than in normal tissues. The 5-gene signature prognostic stratification system based on invasion-related genes could be used to assess prognostic risk in patients with LUAD.     

Identification of a panel of mitotic spindle-related genes as a signature predicting survival in lung adenocarcinoma

Lung adenocarcinoma (LUAD) is one of the most malignant tumor types worldwide. Our objective was to identify a genetic signature that could predict the prognosis of patients with LUAD. We extracted gene data sets from The Cancer Genome Atlas and obtained differentially expressed genes that were highly expressed at every stage. These genes were analyzed using gene set enrichment analysis to obtain four biological processes associated with LUAD. Subsequently, Cox univariate and multivariate analyses were performed to generate four optimized models (G2M checkpoint, E2F targets, mitotic spindle, and glycolysis). We identified a mitotic spindle-related signature (KIF15, BUB1, CCNB2, CDK1, KIF4A, DLGAP5, ECT2, and ANLN), which could be an independent prognostic indicator, to predict the prognosis of patients with LUAD. This new discovery should offer opportunities to explore the pathogenesis of LUAD and prove clinically useful in predicting LUAD patient prognosis.     

Identification of key genes and pathways associated with different immune statuses of hepatitis B virus infection

We aimed to identify key genes and pathways associated with different immune statuses of hepatitis B virus (HBV) infection. The gene expression and DNA methylation profiles were analysed in different immune statuses of HBV infection. Differentially expressed genes (DEGs) and differentially methylated genes (DMGs) were identified, followed by their functional and integrative analyses. The differential expression of IgG Fc receptors (FcγRs) in chronic HBV‐infected patients and immune cells during different stages of HBV infection was investigated. Toll‐like receptor (TLR) signalling pathway (including TLR6) and leucocyte transendothelial migration pathway (including integrin subunit beta 1) were enriched during acute infection. Key DEGs, such as FcγR Ib and FcγR Ia, and interferon‐alpha inducible protein 27 showed correlation with alanine aminotransferase levels, and they were differentially expressed between acute and immune‐tolerant phases and between immune‐tolerant and immune‐clearance phases. The integrative analysis of DNA methylation profile showed that lowly methylated and highly expressed genes, including cytotoxic T lymphocyte‐associated protein 4 and mitogen‐activated protein kinase 3 were enriched in T cell receptor signalling pathway during acute infection. Highly methylated and lowly expressed genes, such as Ras association domain family member 1 and cyclin‐dependent kinase inhibitor 2A were identified in chronic infection. Furthermore, differentially expressed FcγR Ia, FcγR IIa and FcγR IIb, CD3^?CD56⁺CD16⁺ natural killer cells and CD14^highCD16⁺ monocytes were identified between immune‐tolerant and immune‐clearance phases by experimental validation. The above genes and pathways may be used to distinguish different immune statuses of HBV infection.     

A dictionary on microRNAs and their putative target pathways

While in the last decade mRNA expression profiling was among the most popular research areas, over the past years the study of non-coding RNAs, especially microRNAs (miRNAs), has gained increasing interest. For almost 900 known human miRNAs hundreds of pretended targets are known. However, there is only limited knowledge about putative systemic effects of changes in the expression of miRNAs and their regulatory influence. We determined for each known miRNA the biochemical pathways in the KEGG and TRANSPATH database and the Gene Ontology categories that are enriched with respect to its target genes. We refer to these pathways and categories as target pathways of the corresponding miRNA. Investigating target pathways of miRNAs we found a strong relation to disease-related regulatory pathways, including mitogen-activated protein kinase (MAPK) signaling cascade, Transforming growth factor (TGF)-beta signaling pathway or the p53 network. Performing a sophisticated analysis of differentially expressed genes of 13 cancer data sets extracted from gene expression omnibus (GEO) showed that targets of specific miRNAs were significantly deregulated in these sets. The respective miRNA target analysis is also a novel part of our gene set analysis pipeline GeneTrail. Our study represents a comprehensive theoretical analysis of the relationship between miRNAs and their predicted target pathways. Our target pathways analysis provides a ‘miRNA-target pathway’ dictionary, which enables researchers to identify target pathways of differentially regulated miRNAs.     

MicroRNA Alternations in the Testes Related to the Sterility of Triploid Fish

The knowledge of understanding the molecular traits of the sterile triploid fish is sparse. Herein, we analyzed the microRNA (miRNA) alternations in the testes of the sterile triploid fish produced by crossing the tetraploid fish with the diploid fish, compared with those of tetraploids and diploids used as the controls. A total of 136, 134, and 142 conserved miRNAs and 105, 112, and 119 novel miRNAs were identified in the diploid, triploid, and tetraploid fish, respectively. The genes targeted by the differentially expressed miRNAs were identified and were enriched in the GO term cell surface receptor signaling pathway, cellular process, G-protein coupled receptor signaling pathway, and metabolic process. KEGG pathway enrichment was also assessed to evaluate the target genes with differentially expressed miRNAs and these genes were enriched in four pathways (synthesis and degradation of ketone bodies, pentose and glucuronate interconversions, cyanoamino acid metabolic process, and ascorbate and aldarate metabolism). Nine differentially expressed miRNAs were verified by quantitative real-time PCR analysis (qPCR). The upregulated miRNAs in triploids, including miR-101a, miR-199-5p, miR-214, miR-222, and miR-193a, showed the same results with high-throughput sequencing. Among the selected downregulated miRNAs, miR-7b and miR-153b had significantly lower expression levels in triploids. Dnah3 and Tekt1 genes targeted by miR-199-5p showed lower expression in triploids by qPCR. These verified differentially expressed miRNAs may participate in testicular development and sperm activity by targeting functional genes, which were identified with differential expression in the triploid. This evidence provides insights into the epigenetic regulatory mechanisms of sterility in triploid cyprinids.     

Differential expression of microRNAs associated with neurodegenerative diseases and diabetic nephropathy in protein l-isoaspartyl methyltransferase-deficient mice

Protein l -isoaspartyl methyltransferase (PIMT/PCMT1), an enzyme repairing isoaspartate residues in peptides and proteins that result from the spontaneous decomposition of normal l -aspartyl and l -asparaginyl residues during aging, has been revealed to be involved in neurodegenerative diseases (NDDs) and diabetes. However, the molecular mechanisms for a putative association of PIMT dysfunction with these diseases have not been clarified. Our study aimed to identify differentially expressed microRNAs (miRNAs) in the brain and kidneys of PIMT-deficient mice and uncover the epigenetic mechanism of PIMT-involved NDDs and diabetic nephropathy (DN). Differentially expressed miRNAs by sequencing underwent target prediction and enrichment analysis in the brain and kidney of PIMT knockout (KO) mice and age-matched wild-type (WT) littermates. Sequence analysis revealed 40 differentially expressed miRNAs in the PIMT KO mouse brain including 25 upregulated miRNAs and 15 downregulated miRNAs. In the PIMT KO mouse kidney, there were 80 differentially expressed miRNAs including 40 upregulated miRNAs and 40 downregulated miRNAs. Enrichment analysis and a systematic literature review of differentially expressed miRNAs indicated the involvement of PIMT deficiency in the pathogenesis in NDDs and DN. Some overlapped differentially expressed miRNAs between the brain and kidney were quantitatively assessed in the brain, kidney, and serum-derived exosomes, respectively. Despite being preliminary, these results may aid in investigating the pathological hallmarks and identify the potential therapeutic targets and biomarkers for PIMT dysfunction-related NDDs and DN.     

EA15, MIR22, LINC00472 as diagnostic markers for diabetic kidney disease

This study aimed to investigate the molecular mechanisms of diabetic kidney disease (DKD) and to explore new potential therapeutic strategies and biomarkers for DKD. First we analyzed the differentially expressed changes between patients with DKD and the control group using the chip data in Gene Expression Omnibus (GEO) database. Then the gene chip was subjected to be annotated again, so as to screen long noncoding RNAs (lncRNAs) and study expression differences of these lncRNAs in DKD and controlled samples. At last, the function of the differential lncRNAs was analyzed. A total of 252 lncRNAs were identified, and 14 were differentially expressed. In addition, there were 1,629 differentially expressed messenger RNAs (mRNAs) genes, and proliferation and apoptosis adapter protein 15 (PEA15), MIR22, and long intergenic nonprotein coding RNA 472 ( LINC00472) were significantly differentially expressed in DKD samples. Through functional analysis of the encoding genes coexpressed by the three lncRNAs, we found these genes were mainly enriched in type 1 diabetes and autoimmune thyroid disease pathways, whereas in Gene Ontology (GO) function classification, they were also mainly enriched in the immune response, type I interferon signaling pathways, interferon-γ mediated signaling pathways, and so forth. To summary, we identified EA15, MIR22, and LINC00472 may serve as the potential diagnostic markers of DKD.