three-differential-expression-analysis-methods-for-rna-sequencing

Three Differential Expression Analysis Methods for RNA Sequencing: limma, EdgeR, DESeq2

RNA sequencing (RNA-seq) is one of the most widely used technologies in transcriptomics as it can reveal the relationship between the genetic alteration ...

Renmin hospital of Wuhan university

Ovarian cancer (OC), a representative female reproductive system tumor, is one of the most malignant tumors in female. The most important reason for its poor prognosis is because of its high rate of chemotherapy resistance.

miR-21 Induces Chemoresistance in Ovarian Cancer Cells via Mediating the Expression and Interaction of CD44v6 and P-gp.

 Cervical carcinoma is one of the most common malignant gynecological tumors and is associated with high rates of morbidity and mortality. Early diagnosis and early treatment can reduce the mortality rate of cervical cancer. However, there is still no specific biomarkers for the diagnosis and detection of cervical cancer prognosis. Therefore, it is greatly urgent in searching biomarkers correlated with the diagnosis and prognosis of cervical cancer.  The mRNA and microRNA expression profile datasets (GSE7803, GSE9750, GSE63514, and GSE30656) were downloaded from the Gene Expression Omnibus database (GEO). The three microarray datasets were integrated to one via integrated bioinformatics. Differentially expressed genes (DEGs) and microRNAs (DEMs) were obtained by R software. The protein-protein interaction (PPI) networks of the DEGs were performed from the STRING database and further visualized by Cytoscape software. A total of 83 DEGs and 14 DEMs were screened from the microarray expression profile datasets. The miRNAs validated to be associated with cervical cancer were obtained using HMDD online website and the target genes of DEMs were identified using the miRWalk2.0 online database. , and  were the gene targets of hsa-miR-21; the targets of hsa-miR-16 were , and . These targets were all downregulated in cervical cancer. Finally, we verified the expression of those targets in cervical tissues from TCGA and GTEx databases and analyzed their relationship with survival of cervical cancer patients. In the end, the expression of key genes in cervical cancer tissues was verified via experiment method, we found  and  were downregulated in tumor tissues.  This study indicates that  and  are downregulated by the upregulated miR21 and miRNA16 in cervical cancer, respectively, using bioinformatics analysis, and the lower expression of  and  is closely related to the poor prognosis. They might be of clinical significance for the diagnosis and prognosis of cervical cancer, and provide effective targets for the treatment of cervical cancer.

Identification of candidate biomarkers correlated with the diagnosis and prognosis of cervical cancer via integrated bioinformatics analysis.

In this study, we attempted to evaluate the prognostic value of infiltrating immune/stromal cells in clear cell renal cell carcinoma (ccRCC), by using the immune scores and stromal scores based on the "Estimation of STromal and Immune cells in MAlignant Tumours using Expression data" algorithm to represent the levels of infiltrating immune cells and stromal cells. We found that the infiltrating immune cells were associated with poor prognosis of ccRCC. To assess the role of infiltrating immune cells in ccRCC cells, first, we performed differentially expressed genes analysis and functional analysis for validation. The results showed that the underlying mechanism by which infiltrating immune cells promoted cancer progression involved in regulating the nuclear division, angiogenesis, and immune response. Next, we investigated the relationship between infiltrating immune cells and mutations in ccRCC cells. We found that the infiltrating immune cells have certain effects on genetic mutations. In conclusion, infiltrating immune cells within the tumor microenvironment can be used to predict prognosis in ccRCC.

Prognostic value of infiltrating immune cells in clear cell renal cell carcinoma (ccRCC).

Insufficient trophoblast invasion is the key factor for the occurrence of recurrent spontaneous abortions (RSA). Our previous studies identified Yin Yang 1 (YY1) as a transcription factor involved in the regulation of trophoblast invasiveness at the maternal-fetal interface. Long noncoding RNAs (lncRNAs) can regulate gene expression and autophagy in many ways. The purpose of this study was to explore the relationship between YY1 and lncRNAs and the mechanism by which lncRNAs affect the biological behavior of trophoblasts. Bioinformatic analysis predicted that YY1 had three binding sites in the plasmacytoma variant translocation 1 (PVT1) promoter region. Chromatin immunoprecipitation experiments and electrophoretic mobility shift assays verified that YY1 can directly bind to the PVT1 promoter. Compared with its expression levels in human placental villi tissue samples from the normal pregnancy group, the PVT1 expression levels were significantly lower in tissues from the RSA group. PVT1 knockdown significantly reduced adhesion, invasion, autophagy, and mTOR expression in HTR-8/SVneo cells and greatly increased apoptosis in vitro. This study revealed a novel regulatory pathway in which YY1 can act directly on PVT1 promoter to regulate its transcription, which further affects trophoblast invasion and adhesion by regulating autophagy via the mTOR pathway, and these effects might be involved in RSA pathogenesis.

YY1-PVT1 affects trophoblast invasion and adhesion by regulating mTOR pathway-mediated autophagy.

 Cisplatin (CDDP) is a widely used and effective basic chemotherapeutic drug for the treatment of a variety of tumors, including ovarian cancer. However, adverse side effects and acquired drug resistance are observed in the clinical application of CDDP. Identifying a mode of administration that can alleviate side effects and reduce drug resistance has become a promising strategy to solve this problem.  In this study, 3D printing technology was used to prepare a CDDP-poly (lactic-co-glycolic acid) (CDDP-PLGA) polymer compound stent, and its physicochemical properties and cytotoxicity were evaluated both  and .  The CDDP-PLGA stent had a significant effect on cell proliferation and apoptosis and clearly decreased the size of subcutaneous tumors in nude mice, whereas the systemic side effects were mild compared with those of intraperitoneal CDDP injection. Compared with the control group, CDDP-PLGA significantly increased the mRNA and protein levels of p-glycoprotein ( < 0.01;  < 0.01) and decreased vascular endothelial growth factor mRNA ( < 0.05) and protein levels ( < 0.01), however, CDDP-PLGA significantly decreased the mRNA and protein levels of p-glycoprotein ( < 0.01;  < 0.01) and vascular endothelial growth factor ( < 0.01;  < 0.01), which are associated with chemoresistance, in subcutaneous tumor tissue. Immunohistochemistry assay results revealed that, in the CDDP-PLGA group, the staining of the proliferation-related genes Ki67 and PCNA were lightly, and the apoptosis-related gene caspase-3 stained deeply.  PLGA biomaterials loaded with CDDP, as compared with the same amount of free CDDP, showed good efficacy in terms of cytotoxicity, as evidenced by changes in apoptosis. Continuous local CDDP release can decrease the systemic side effects of this drug and the occurrence of drug resistance and angiogenesis, and improve the therapeutic effect. This new approach may be an effective strategy for the local treatment of epithelial ovarian cancer.

The role of a drug-loaded poly (lactic co-glycolic acid) (PLGA) copolymer stent in the treatment of ovarian cancer.

Polycystic ovary syndrome (PCOS) is a life-long reproductive, endocrine, and metabolic disorder that affects up to 17% of women of reproductive age. However, the effect of granulosa cells (GCs) transcriptome changes on oocyte capacity and follicular development in patients with PCOS has not been elucidated. This study aims to analyze transcriptome changes in GCs of PCOS from different perspectives and explore potential biomarkers for the diagnosis and treatment of PCOS. The gene expression profiles of GSE34526 and GSE107746 were obtained from the GEO database. Differentially expressed genes (DEGs) and key signaling pathways were identified. Gene Set Enrichment Analysis (GSEA) revealed that Toll-like receptors, NOD-like receptors, and NOTCH signaling pathways were obviously enriched in GCs of PCOS. We further analyzed DEGs from three aspects: transcription factors (TFs), secreted proteins, and follicular development. Compared with normal GCs, the DEGs encoding TFs and secretory proteins in GCs of PCOS remarkably changed. Besides, HAS2 and CBLN1, which are highly expressed in preovulatory follicular GCs and may trigger ovulation, were significantly decreased in GCs of PCOS. This study found candidate genes and signaling pathways in PCOS, providing new insights and foundations for the etiology of PCOS. Besides, HSA2 and CBLN1 may be potential therapeutic biomarkers for ovulation disorders in PCOS.

Identification of Potential Biomarkers of Polycystic Ovary Syndrome via Integrated Bioinformatics Analysis.

Cytotoxic T-lymphocyte associated protein 4 (CTLA4) inhibitors have been shown to significantly prolong the overall survival (OS) in a wide range of cancers. However, its application in clear cell renal cell carcinoma (ccRCC) is limited due to the therapy response, and the prognostic value of CTLA4 in ccRCC has not been investigated in detail.

CTLA4 has a profound impact on the landscape of tumor-infiltrating lymphocytes with a high prognosis value in clear cell renal cell carcinoma (ccRCC).

Pelvic organ prolapses (POP) notably reduces the quality of life in elderly populations due to bladder and bowel dysfunction, incontinence, and coital problems. Extracellular matrix (ECM) disorder is a pivotal event in the progression of POP, but to date, its specific underlying mechanism remains unclear. The ligaments of patients with POP and healthy controls were collected to compare the expression of Homeobox11 (HOXA11) and transforming growth factor β (TGF‑β1) via immunohistochemical analysis. HOXA11 and TGF‑β1 were overexpressed or knocked down in fibroblast cells to explore their effects on the expression of collagen and matrix metalloproteinases (MMPs). HOXA11 and TGF‑β1 were greatly reduced in the ligaments of patients with POP. The overexpression and downregulation of HOXA11 and TGF‑β1 can mediate ECM disorder via regulating expression of collagen (Col) and MMPs. In addition, HOXA11 and TGF‑β1 exerted synergistic effect on the expression of Col and MMPs. The present study identified that HOXA11 and TGF‑β1 serve critical roles in mediating ECM disorders, which may be of clinical significance for the diagnosis and treatment of patients with POP.

Molecular mechanism of extracellular matrix disorder in pelvic organ prolapses.

Subsequently to the publication of the above article, the authors have realized that the bar charts shown for Fig. 3A and B, as they appeared in the paper, were the same as the bar charts shown for Fig. 4B and D. Fig. 3, as it should have appeared, is shown below. All the authors agree to this Corrigendum. Note that the revisions made to this figure do not adversely affect the results reported in the paper, or the conclusions stated therein. The authors regret that the duplication of the histograms in Fig. 4 as Fig. 3 was not noticed prior to the publication of this article, and offer their apologies to the Editor of Molecular Medicine Reports and to the readers of the Journal. [the original article was published in Molecular Medicine Reports 22: 4611-4618, 2020; DOI: 10.3892/mmr.2020.11564].

[Corrigendum] Molecular mechanism of extracellular matrix disorder in pelvic organ prolapses.

Hepatocellular carcinoma (HCC) ranks fifth among common cancers and is the second most common cause of cancer-related mortality worldwide. This study is aimed at identifying an immune-related long noncoding RNA (lncRNA) signature as a potential biomarker with prognostic value to improve early diagnosis and provide potential therapeutic targets for HCC patients. The subjects of this study were HCC samples with complete transcriptome data and clinical information downloaded from The Cancer Genome Atlas (TCGA) database. We then extracted the immune-related mRNA and lncRNA expression profiles. Based on the expression profiles of immune-related lncRNAs, we identified a nine-lncRNA signature that was related to the progression of HCC. The risk score was calculated based on the expression level of the nine lncRNAs of each sample, which divided patients into high-risk and low-risk groups. We found that the increased risk score was associated with a poor prognosis of HCC patients. To assess the accuracy of the survival model, we calculated a receiver operating characteristic (ROC) for validation. The curve showed that the area under the curve (AUC) for the risk score was 0.792. Besides, both principal component analysis (PCA) and gene set enrichment analysis (GSEA) were further used for functional annotation. We found that the distribution patterns were different between the low-risk and high-risk groups in PCA, and the underlying mechanism by which the nine lncRNAs promoted the progression of HCC involved an abnormal immune status. Finally, we analyzed the infiltration of twenty-nine kinds of immune cells and the activation of immune function in HCC using the ssGSEA algorithm. The results showed that aDCs, iDCs, macrophages, Tfh, Th1, Treg, and NK cells were correlated with the progress of HCC patients. And the immune functions including APC costimulation, CCR, check point, HLA, MHC class I, and Type II IFN responses were also significantly different between the high-risk and low-risk groups. In conclusion, our study identified a nine-lncRNA signature with potential prognostic value for patients with HCC, which could be used as a new biomarker for the diagnosis and immunotherapy of HCC.

Identification of a Nine Immune-Related lncRNA Signature as a Novel Diagnostic Biomarker for Hepatocellular Carcinoma.

Polycystic ovary syndrome (PCOS) is the most common hormonal disorder among reproductive-aged women worldwide, however, the mechanisms and progression of PCOS still unclear due to its heterogeneous nature. Using the human granulosa-like tumor cell line (KGN) and PCOS mice model, we explored the function of lncRNA UCA1 in the pathological progression of PCOS.

Silencing of lncRNA UCA1 inhibited the pathological progression in PCOS mice through the regulation of PI3K/AKT signaling pathway.

Hepatocellular carcinoma (HCC) is one of the main causes of cancer-associated deaths globally, accounts for 90% of primary liver cancers. However, further studies are needed to confirm the metabolism-related gene signature related to the prognosis of patients with HCC.

Shiyi Liu

Department of Obstetrics and Gynecology

Renmin hospital of Wuhan university

Three Differential Expression Analysis Methods for RNA Sequencing: limma, EdgeR, DESeq2

Shiyi Liu

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Obstetrics and Gynecology

Renmin hospital of Wuhan university

Three Differential Expression Analysis Methods for RNA Sequencing: limma, EdgeR, DESeq2

Department of Obstetrics and Gynecology