MiRNA expression of tube forming HUVECs
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE56663 | GEO | 2017/08/11
SECONDARY ACCESSION(S): PRJNA244256
REPOSITORIES: GEO
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2014-02-20 | E-GEOD-44597 | biostudies-arrayexpress
Project description:Sparstolonin B is a novel bioactive compound isolated from Sparganium stoloniferum, an herb historically used in Traditional Chinese Medicine as an anti-tumor agent. SsnB has previously demonstrated anti-angiogenic properties. In functional assays, SsnB inhibited endothelial cell tube formation (Matrigel method) and cell migration (Transwell method) in a dose-dependent manner. We used microarrays to examine how SsnB affected the gene expression of human umbilical vein endothelial cells (HUVECs), focusing in particular on pathways related to angiogenesis.
2014-02-20 | GSE44597 | GEO
Project description:Kallikrein-related peptidase 3 (KLK3, also known as prostate-specific antigen, PSA) is a chymotrypsin-like kallikrein that has been shown to have anti-angiogenic properties. KLK3 expression in prostate cancer is associated with low microvessel density. We have previously shown in a human umbilical vein endothelial cell (HUVEC) model that the anti-angiogenic effect of KLK3 is related to its enzymatic activity. However, the mechanism of this effect remains to be clarified. To this end we have used a DNA microarray to study the KLK3-induced changes in gene expression associated with reduction of HUVEC tube formation. Among the 41000 genes studied, 311 were differentially expressed in control and KLK3-treated cells. These changes were enriched in several pathways, including the pathways associated with proteasome, ubiquitin-mediated proteolysis, focal adhesion and regulation of actin cytoskeleton. Furthermore, the changes were opposite to those described previously to occur during tubulogenesis. In conclusion, our results show that KLK3 induces gene expression changes in HUVECs. While these changes might be relevant for the mechanism by which KLK3 exerts its anti-angiogenic activity, it cannot be judged by the present results whether the changes are secondary to morphogenic differentiation of the cells or the primary mechanism mediating the effect of KLK3. Keywords: KLK3 treatment, tube formation Human umbilical vein endothelial cells (HUVECs) were isolated from umbilical cord veins and grown in Endothelial Cell Basal Medium supplemented with an Endothelial Cell Growth Medium Supplement Pack (PromoCell). The cells were cultured on Matrigel basement membrane prepara-tion together with 500 nM enzymatically active KLK3 (PSA). Phosphate buffered saline (PBS) was used for control. HUVECs were grown on Matrigel for 18 h. RNA was isolated from KLK3-treated and control HUVECs from three different individuals. Cell Recovery Solution (BD Biosciences) was used to detach cells from the Matrigel. Total RNA was isolated with the RNeasy Mini Kit (Qiagen) according to manufacturerâ??s instructions. RNA con-centration and quality of the samples was determined with an Agilent 2100 bioanalyser (Agilent Technologies) by measuring the RNA integrity number (RIN). All samples contained high quality RNA (RIN 9.5-10). DNA microarray analysis was performed with the Agilent Whole Human Genome Oligo Microar-ray 4x44K (G4112F) (Agilent Technologies Inc.) following the manufacturerâ??s protocol. Total RNA (1 ï?g) was labeled using the RNA input fluorescent linear amplification kit according to the manufacturer's recommendations (Agilent Technologies Inc.). RNA was hybridized on Agilent's Human 4x44K Oligo Microarrays containing 44000 features (41000 unique genes and transcripts). Pre-processing of the data was performed by Feature Extraction software (v 7.5.1).
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Project description:BACKGROUND: Development of the neural tube is a highly orchestrated process relying on precise, spatio-temporal expression of numerous genes as well as hierarchies of signal transduction and gene regulatory networks. Disruption of expression of a number of genes participating in these networks is believed to underlie developmental anomalies such as neural tube defects (NTDs) resulting from anomalous neural tube morphogenesis. MicroRNAs (miRNAs), a large family of noncoding RNAs, have been shown to function as gene silencers, and thus, are key modulators of cell and tissue differentiation. To elucidate potential roles of miRNAs in murine neural tube development, miRNA gene expression profiling has been utilized in the current study, to garner novel and in-depth knowledge on the expression and regulation of genes encoding miRNAs as well as their potential target genes governing maturation of the mammalian neural tube. METHODS: With the aim of identifying differentially expressed miRNAs during mammalian neural tube ontogenesis, miRNA expression profiles from gestation day (GD) -8.5, -9.0 and -9.5 murine neural tube tissue were compared utilizing miRXplore™ microarrays from Miltenyi Biotech GmbH. Gene expression changes observed in microarray analysis were verified by TaqManTM quantitative Real-Time PCR. clValid R package and the UPGMA (hierarchical) clustering method were utilized for cluster analysis of the microarray data. Functional associations among selected miRNAs were examined exploiting Ingenuity Pathway Analysis. RESULTS: Expression of approximately 12% of the 609 murine miRNA genes examined was detected in murine neural tube tissues from GD -8.5, -9.0 and -9.5. Clustering analysis revealed several developmentally regulated clusters among these expressed genes. MicroRNA target analysis enabled identification of a panoply of protein-coding target genes of the differentially expressed miRNAs within such clusters. Interestingly, many of these target genes have been shown to be associated with vital cellular processes such as cell proliferation, cell adhesion, cell migration, differentiation, apoptosis and epithelial-mesenchymal transformation, all of which are essential for normal neural tube development. Utilization of Ingenuity Pathway Analysis (IPA; Ingenuity Systems) allowed identification of interactive biological networks connecting differentially expressed miRNAs and their target genes highlighting functional relationships. CONCLUSIONS: In the present study, a unique gene expression signature of a range of miRNAs in embryonic neural tube tissue was delineated. Analysis of miRNA target genes and gene interaction pathways emphasized that expression of numerous protein-encoding genes, indispensable for normal neural tube morphogenesis, may be regulated by specific miRNAs.
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