Browsing by Subject "Gene Expression Profiling"
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Item A Comparison of Gene Expression Profiles between Glucocorticoid Responder and Non-Responder Bovine Trabecular Meshwork Cells Using RNA Sequencing(PLOS, 2017-01-09) Bermudez, Jaclyn Y.; Webber, Hannah C.; Brown, Bartley; Braun, Terry A.; Clark, Abbot F.; Mao, WeimingThe most common ocular side effect of glucocorticoid (GC) therapy is GC-induced ocular hypertension (OHT) and GC-induced glaucoma (GIG). GC-induced OHT occurs in about 40% of the general population, while the other 60% are resistant. This study aims to determine the genes and pathways involved in differential GC responsiveness in the trabecular meshwork (TM). Using paired bovine eyes, one eye was perfusion-cultured with 100nM dexamethasone (DEX), while the fellow eye was used to establish a bovine TM (BTM) cell strain. Based on maximum IOP change in the perfused eye, the BTM cell strain was identified as a DEX-responder or non-responder strain. Three responder and three non-responder BTM cell strains were cultured, treated with 0.1% ethanol or 100nM DEX for 7 days. RNA and proteins were extracted for RNA sequencing (RNAseq), qPCR, and Western immunoblotting (WB), respectively. Data were analyzed using the human and bovine genome databases as well as Tophat2 software. Genes were grouped and compared using Student's t-test. We found that DEX induced fibronectin expression in responder BTM cells but not in non-responder cells using WB. RNAseq showed between 93 and 606 differentially expressed genes in different expression groups between responder and non-responder BTM cells. The data generated by RNAseq were validated using qPCR. Pathway analyses showed 35 pathways associated with differentially expressed genes. These genes and pathways may play important roles in GC-induced OHT and will help us to better understand differential ocular responsiveness to GCs.Item Identification of long non-coding RNA-related and -coexpressed mRNA biomarkers for hepatocellular carcinoma(BioMed Central Ltd., 2019-01-31) Zhang, Fan; Ding, Linda; Cui, Li; Barber, Robert C.; Deng, BinBackground: While changes in mRNA expression during tumorigenesis have been used widely as molecular biomarkers for the diagnosis of a number of cancers, the approach has limitations. For example, traditional methods do not consider the regulatory and positional relationship between mRNA and lncRNA. The latter has been largely shown to possess tumor suppressive or oncogenic properties. The combined analysis of mRNA and lncRNA is likely to facilitate the identification of biomarkers with higher confidence. Results: Therefore, we have developed an lncRNA-related method to identify traditional mRNA biomarkers. First we identified mRNAs that are differentially expressed in Hepatocellular Carcinoma (HCC) by comparing cancer and matched adjacent non-tumorous liver tissues. Then, we performed mRNA-lncRNA relationship and coexpression analysis and obtained 41 lncRNA-related and -coexpressed mRNA biomarkers. Next, we performed network analysis, gene ontology analysis and pathway analysis to unravel the functional roles and molecular mechanisms of these lncRNA-related and -coexpressed mRNA biomarkers. Finally, we validated the prediction and performance of the 41 lncRNA-related and -coexpressed mRNA biomarkers using Support Vector Machine model with five-fold cross-validation in an independent HCC dataset from RNA-seq. Conclusions: Our results suggested that mRNAs expression profiles coexpressed with positionally related lncRNAs can provide important insights into early diagnosis and specific targeted gene therapy of HCC.Item Mirna Expression in Glaucomatous and TGFbeta2 Treated Lamina Cribrosa Cells(MDPI, 2021-06-08) Lopez, Navita N.; Rangan, Rajiv; Clark, Abbot F.; Tovar-Vidales, TaraGlaucoma is a group of optic neuropathies that leads to irreversible vision loss. The optic nerve head (ONH) is the site of initial optic nerve damage in glaucoma. ONH-derived lamina cribrosa (LC) cells synthesize extracellular matrix (ECM) proteins; however, these cells are adversely affected in glaucoma and cause detrimental changes to the ONH. LC cells respond to mechanical strain by increasing the profibrotic cytokine transforming growth factor-beta 2 (TGFbeta2) and ECM proteins. Moreover, microRNAs (miRNAs or miR) regulate ECM gene expression in different fibrotic diseases, including glaucoma. A delicate homeostatic balance between profibrotic and anti-fibrotic miRNAs may contribute to the remodeling of ONH. This study aimed to determine whether modulation of miRNAs alters the expression of ECM in human LC cells. Primary human normal and glaucoma LC cells were grown to confluency and treated with or without TGFbeta2 for 24 h. Differences in expression of miRNAs were analyzed using miRNA qPCR arrays. miRNA PCR arrays showed that the miR-29 family was significantly decreased in glaucomatous LC cell strains compared to age-matched controls. TGFbeta2 treatment downregulated the expression of multiple miRNAs, including miR-29c-3p, compared to controls in LC cells. LC cells transfected with miR-29c-3p mimics or inhibitors modulated collagen expression.Item Stress and interferon signalling-mediated apoptosis contributes to pleiotropic anticancer responses induced by targeting NGLY1(Springer Nature, 2018-11-02) Zolekar, Ashwini; Lin, Victor J. T.; Mishra, Nigam M.; Ho, Yin Ying; Hayatshahi, Hamed S.; Parab, Abhishek; Sampat, Rohit; Liao, Xiaoyan; Hoffmann, Peter; Liu, Jin; Emmitte, Kyle A.; Wang, Yu-ChiehBACKGROUND: Although NGLY1 is known as a pivotal enzyme that catalyses the deglycosylation of denatured glycoproteins, information regarding the responses of human cancer and normal cells to NGLY1 suppression is limited. METHODS: We examined how NGLY1 expression affects viability, tumour growth, and responses to therapeutic agents in melanoma cells and an animal model. Molecular mechanisms contributing to NGLY1 suppression-induced anticancer responses were revealed by systems biology and chemical biology studies. Using computational and medicinal chemistry-assisted approaches, we established novel NGLY1-inhibitory small molecules. RESULTS: Compared with normal cells, NGLY1 was upregulated in melanoma cell lines and patient tumours. NGLY1 knockdown caused melanoma cell death and tumour growth retardation. Targeting NGLY1 induced pleiotropic responses, predominantly stress signalling-associated apoptosis and cytokine surges, which synergise with the anti-melanoma activity of chemotherapy and targeted therapy agents. Pharmacological and molecular biology tools that inactivate NGLY1 elicited highly similar responses in melanoma cells. Unlike normal cells, melanoma cells presented distinct responses and high vulnerability to NGLY1 suppression. CONCLUSION: Our work demonstrated the significance of NGLY1 in melanoma cells, provided mechanistic insights into how NGLY1 inactivation leads to eradication of melanoma with limited impact on normal cells, and suggested that targeting NGLY1 represents a novel anti-melanoma strategy.