Analysis of microRNAs expressions in chondrosarcoma

Abstract

MicroRNAs (miRNAs) are small non-coding RNAs capable of inhibiting gene expression post-transcriptionally and expression profiling can provide therapeutic targets and tools for cancer diagnosis. Chondrosarcoma is a mesenchymal tumor with unknown cause and differentiation status. Here, we profiled miRNA expression of chondrosarcoma, namely clinical samples from human conventional chondrosarcoma tissue, established chondrosarcoma cell lines, and primary non-tumorous adult articular chondrocytes, by miRNA array and quantitative real-time PCR. A wide variety of miRNAs were differently downregulated in chondrosarcoma compared to non-tumorous articular chondrocytes; 27 miRNAs: miR-10b, 23b, 24-1*, 27b, 100, 134, 136, 136*, 138, 181d, 186, 193b, 221*, 222, 335, 337-5p, 376a, 376a*, 376b, 376c, 377, 454, 495, 497, 505, 574-3p, and 660, were significantly downregulated in chondrosarcoma and only 2: miR-96 and 183, were upregulated. We further validated the expression levels of miRNAs by quantitative real-time PCR for miR-181a, let-7a, 100, 222, 136, 376a, and 335 in extended number of chondrosarcoma clinical samples. Among them, all except miR-181a were found to be significantly downregulated in chondrosarcoma derived samples. The findings provide potential diagnostic value and new molecular understanding of chondrosarcoma.

Keywords: cartilage; chondrosarcoma; malignancy; microRNA; sarcoma.

Figure 1

Correlations between samples on the miRNA arrays. (a) Correlation matrix for log-intensity among samples. (b) Hierarchical clustering of the arrays. The dendrogram shows the degree of similarity of miRNA expression pattern among samples. The chondrogenic samples, including both chondrosarcoma clinical samples and chondrosarcoma derived cell lines, showed a distinct expression pattern in miRNA from primary non-tumorous chondrocyte derived samples. CS_clin = chondrosarcoma clinical sample; CS_cell = chondrosarcoma derived cell line; PNC = primary non-tumorous articular chondrocytes.

Figure 2

Expression pattern of the selected 117 miRNAs. The closed triangles show the miRNAs that were subjected to the further verification by quantitative real-time PCR.

Figure 3

Differentially expressed miRNAs in chondrogenic samples. The expression levels of miRNAs in clinical chondrosarcoma samples and chondrosarcoma cell line samples were compared to primary non-tumorous articular chondrocytes. (a) The list of upregulated miRNAs in chondrosarcomas vs PNC (b) The list of downregulated miRNAs in chondrosarcomas vs PNC. The underlined miRNAs represent those used for verification by quantitative real-time PCR. CS_clin = chondrosarcoma clinical sample; CS_cell = chondrosarcoma derived cell line; PNC = primary non-tumorous articular chondrocytes.

Figure 4

Expression of miRNAs in chondrosarcoma clinical samples in quantitative real-time PCR assay. All data are given as fold change expression levels compared to PNC. The expression level of each miRNA was normalized to RNU6B as an internal control. CS_clin = chondrosarcoma clinical sample; CS_cell = chondrosarcoma derived cell line; PNC = primary non-tumorous articular chondrocytes. The bar represents the mean. *p < .05 by ANOVA and pairwise t-test with Bonferroni's correction.