Project description:MicroRNA 21 (miR-21) has been implicated in various aspects of carcinogenesis. However, the function and molecular mechanism of miR-21 in cervical squamous carcinoma has not been studied. Using TaqMan quantitative real-time PCR and Northern blot, we confirmed that miR-21 is significantly overexpressed in human cervical squamous cancer tissues and cell lines. Remarkably, we showed that the level of miR-21 correlates with the nodal status and differentiation by ISH. Furthermore, we demonstrated that miR-21 regulates cervical squamous cells proliferation, apoptosis, and migration which are HPV16 positive. In order to identify the candidate target genes for miR-21, we used gene expression profiling. By luciferase reporter assay, we confirmed the CCL20 gene is one of its targets, which is relative to the HPV16 oncogenes E6 and E7. Our results suggest that miR-21 may be involved in the cervical squamous cell tumorigenesis.
Project description:MicroRNA 21 (miR-21) has been implicated in various aspects of carcinogenesis. However, the function and molecular mechanism of miR-21 in cervical squamous carcinoma has not been studied. Using TaqMan quantitative real-time PCR and Northern blot, we confirmed that miR-21 is significantly overexpressed in human cervical squamous cancer tissues and cell lines. Remarkably, we showed that the level of miR-21 correlates with the nodal status and differentiation by ISH. Furthermore, we demonstrated that miR-21 regulates cervical squamous cells proliferation, apoptosis, and migration which are HPV16 positive. In order to identify the candidate target genes for miR-21, we used gene expression profiling. By luciferase reporter assay, we confirmed the CCL20 gene is one of its targets, which is relative to the HPV16 oncogenes E6 and E7. Our results suggest that miR-21 may be involved in the cervical squamous cell tumorigenesis. Total RNA of cells transfected with anti-miR-21 or scrambled RNA oligonucleotide was extracted using the TRIZOL Reagent according to the manufacturer's instructions. Gene-expression profiling was performed for each pooling RNA sample separately on the GeneChip_ Porcine Genome Array (Affymetrix) at CapitalBio Corporation (Beijing, China) in which GeneChip microarray service was certificated by Affymetrix.
Project description:Little is known about the alterations in microRNA (miRNA) expression patterns during the consecutive stages of cervical cancer development and their association with chromosomal instability and epigenetic changes. We integrated miRNA expression profiles in normal cervical squamous epithelium, high-grade precancerous lesions (CIN2-3), squamous cell carcinomas (SCC) and adenocarcinomas (AdCAs) with previously generated chromosomal profiles of the same samples. In addition, the DNA methylation status of downregulated miRNAs located in a CpG island was determined. Significantly differential expression during the consecutive stages of cervical SCC development was observed for 106 miRNAs. Altered expression of 5 significantly differentially expressed miRNAs, hsa-miR-9 (1q23.2), hsa-miR-15b (3q25.32), hsa-miR-28-5p (3q27.3), hsa-miR-100 and hsa-miR-125b (both 11q24.1) was directly linked to frequent chromosomal alterations. Another 9 significantly downregulated miRNAs were located within a CpG island. Three of these 9 miRNAs, hsa-miR-203, hsa-miR-572, and hsa-miR-638, showed increased methylation in cervical cancer cell lines and HPV-immortalised cells compared to primary keratinocytes. Functional analyses were performed for hsa-miR-9, representing a potential oncogene with increased expression linked to a chromosomal gain, and hsa-miR-203, representing a potential tumour suppressor gene silenced by DNA methylation. Hsa-miR-9 and hsa-miR-203 were found to alter cell viability and anchorage independent growth in vitro, respectively, supporting their oncogenic and tumour suppressive function in cervical cancer. In conclusion, differential expression of 106 miRNAs, partly associated with chromosomal alterations and epigenetic changes, was observed during cervical SCC development. Altered expression of hsa-miR-9 and hsa-miR-203 was shown to be functionally relevant, underlining the importance of deregulated miRNA expression in cervical carcinogenesis. 10 squamous cell carcinomas of the cervix, 9 adenocarcinomas of the cervix, 18 high-grade cervical intraepithelial neoplasias (CIN2-3), and 10 cervical squamous epithelial samples with normal histology were analysed using single channel (Cy3) miRNA microarrays from Agilent (G4471A-016436).
Project description:10 normal squamous cervical epitheilia samples, 7 high grade squamous intraepithelial lesions, and 21 invasive squamous cell carcinomas of the cervix samples were obtained using laser capture miicrodissection. Two rounds of T7-based linear RNA amplification using the Arcturus RiboAmp kit were performed for each sample, and assayed using Affymetrix HG_U133A arrays. Experiment Overall Design: 10 normal squamous cervical epitheilia samples, 7 high grade squamous intraepithelial lesions, and 21 invasive squamous cell carcinomas of the cervix, each from different patients, were each assayed on single HG_U133A arrays. Three additional test samples were also assayed. Experiment Overall Design: The log-transformed probe-set values and the results of the statistical analysis for each probe-set, and the associated README file, are included as Supplementary files.
Project description:We sought to apply the technologies of gene expression profiling to detect genes significant in the aetiology of cervical carcinoma . We investigated 14 normal (NAD), 11 low grade squamous intrapepithelial lesions (LSIL), 21 high grade squamous intraepithelial lesions (HSIL) and 28 squamous cell carcinomas by Affymetrix GeneChip whole transcriptome profiling. Two SCC cell lines were also included in the cohort. Normal and SILS were profiled using the Affymetrix U133A platform, while SCCs and Cell lines were profiled using the Affymetrix U133A plus 2.0 array. This submission describes the transcriptional profiles of a cohort totalling 77 cervical normal, premalignant lesions, and squamous cell carcinomas
Project description:10 normal squamous cervical epitheilia samples, 7 high grade squamous intraepithelial lesions, and 21 invasive squamous cell carcinomas of the cervix samples were obtained using laser capture miicrodissection. Two rounds of T7-based linear RNA amplification using the Arcturus RiboAmp kit were performed for each sample, and assayed using Affymetrix HG_U133A arrays. Keywords: disease state analysis
Project description:Little is known about the alterations in microRNA (miRNA) expression patterns during the consecutive stages of cervical cancer development and their association with chromosomal instability and epigenetic changes. We integrated miRNA expression profiles in normal cervical squamous epithelium, high-grade precancerous lesions (CIN2-3), squamous cell carcinomas (SCC) and adenocarcinomas (AdCAs) with previously generated chromosomal profiles of the same samples. In addition, the DNA methylation status of downregulated miRNAs located in a CpG island was determined. Significantly differential expression during the consecutive stages of cervical SCC development was observed for 106 miRNAs. Altered expression of 5 significantly differentially expressed miRNAs, hsa-miR-9 (1q23.2), hsa-miR-15b (3q25.32), hsa-miR-28-5p (3q27.3), hsa-miR-100 and hsa-miR-125b (both 11q24.1) was directly linked to frequent chromosomal alterations. Another 9 significantly downregulated miRNAs were located within a CpG island. Three of these 9 miRNAs, hsa-miR-203, hsa-miR-572, and hsa-miR-638, showed increased methylation in cervical cancer cell lines and HPV-immortalised cells compared to primary keratinocytes. Functional analyses were performed for hsa-miR-9, representing a potential oncogene with increased expression linked to a chromosomal gain, and hsa-miR-203, representing a potential tumour suppressor gene silenced by DNA methylation. Hsa-miR-9 and hsa-miR-203 were found to alter cell viability and anchorage independent growth in vitro, respectively, supporting their oncogenic and tumour suppressive function in cervical cancer. In conclusion, differential expression of 106 miRNAs, partly associated with chromosomal alterations and epigenetic changes, was observed during cervical SCC development. Altered expression of hsa-miR-9 and hsa-miR-203 was shown to be functionally relevant, underlining the importance of deregulated miRNA expression in cervical carcinogenesis.
Project description:We sought to apply the technologies of gene expression profiling to detect genes significant in the aetiology of cervical carcinoma . We investigated 14 normal (NAD), 11 low grade squamous intrapepithelial lesions (LSIL), 21 high grade squamous intraepithelial lesions (HSIL) and 28 squamous cell carcinomas by Affymetrix GeneChip whole transcriptome profiling. Two SCC cell lines were also included in the cohort. Normal and SILS were profiled using the Affymetrix U133A platform, while SCCs and Cell lines were profiled using the Affymetrix U133A plus 2.0 array.
Project description:Cervical cancer is the second most common cancer in women worldwide. In addition to the important role played by HPV, the underlying mechanism promoting cervical tumorigenesis is complex and involves deregulation of key signaling pathways. Recently, role of miRNA mediated abnormal regulatory mechanisms is implicated in the pathogenesis of cervical cancer. Micro RNAs are regulatory, non‐coding RNAs about 21–23 nucleotides in length and effects the expression of a number of genes at the post‐transcriptional level. For the past few decades, role of curcumin in inhibiting the growth of cervical cancer and increasing the chemo and radio- sensistivity has been studied extensively. Interestingly, curcumin was shown to downregulate NF-κB, Wnt/β-catenin, TGF‐β and various other signaling pathways in cervical cancer cells. Although, a number of microarray studies have examined the use of miRNAs as cancer diagnostic markers, the regulation of miRNAs upon treatment with curcumin in cervical cancer cells has not been studied. The current study is aimed to perform miRNA profiling in cervical cancer cells following curcumin treatment and to study the role of miRNAs in regulating the different signaling pathways.