Limits...
MicroRNA expression profiling identifies activated B cell status in chronic lymphocytic leukemia cells.

Li S, Moffett HF, Lu J, Werner L, Zhang H, Ritz J, Neuberg D, Wucherpfennig KW, Brown JR, Novina CD - PLoS ONE (2011)

Bottom Line: Gene set enrichment analysis (GSEA) identified statistically-significant similarities in miRNA expression between activated B cells and CLL cells including upregulation of miR-34a, miR-155, and miR-342-3p and downregulation of miR-103, miR-181a and miR-181b.Additionally, decreased levels of two CLL signature miRNAs miR-29c and miR-223 are associated with ZAP70(+) and IgV(H) unmutated status and with shorter time to first therapy.These data indicate an activated B cell status for CLL cells and suggest that the direction of change of individual miRNAs may predict clinical course in CLL.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, Massachusetts, United States of America.

ABSTRACT
Chronic lymphocytic leukemia (CLL) is thought to be a disease of resting lymphocytes. However, recent data suggest that CLL cells may more closely resemble activated B cells. Using microRNA (miRNA) expression profiling of highly-enriched CLL cells from 38 patients and 9 untransformed B cells from normal donors before acute CpG activation and 5 matched B cells after acute CpG activation, we demonstrate an activated B cell status for CLL. Gene set enrichment analysis (GSEA) identified statistically-significant similarities in miRNA expression between activated B cells and CLL cells including upregulation of miR-34a, miR-155, and miR-342-3p and downregulation of miR-103, miR-181a and miR-181b. Additionally, decreased levels of two CLL signature miRNAs miR-29c and miR-223 are associated with ZAP70(+) and IgV(H) unmutated status and with shorter time to first therapy. These data indicate an activated B cell status for CLL cells and suggest that the direction of change of individual miRNAs may predict clinical course in CLL.

Show MeSH

Related in: MedlinePlus

Validation of altered miRNA expression identified by GSEA.miRNA-specific RT-PCR independently confirms altered expression of GSEA-identified miRNAs in B cells activated with CpG. A. miR-181a, miR-181b, miR-20b, miR-103, miR-15b, miR-331-3p, and let-7c. B. miR-155. C. miR-342-3p, miR-337-3p, miR-198, and miR-34a. For (A, B, C), B (-): control B cell samples. B (+): corresponding activated B samples by CpG. Error bars indicate the standard deviation of 3 independent samples. D, E. miRNA-specific RT-PCR confirms the differential expression of subsets of GSEA-identified miRNAs in CLL relative to control B cells. Upregulated miRNAs are miR-34a, miR-155, miR-198 and miR-342-3p. Downregulated miRNA expression including let-7c, miR-15b, miR-20b, miR-103, miR-181a, miR-181b, miR-331-3p and miR-337-3p. Between 4 to 8 control unstimulated B cell samples (square) and between 6 to 21 CLL patient samples (triangle) were used. The bar indicates the average expression in each group. p-value for the analysis calculated by t-test is shown. miRNA expression was normalized by snoRNA-RNU44 expression. Relative miRNA expression was calculated by ΔΔCt method and is indicated on the Y-axis.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3050979&req=5

pone-0016956-g002: Validation of altered miRNA expression identified by GSEA.miRNA-specific RT-PCR independently confirms altered expression of GSEA-identified miRNAs in B cells activated with CpG. A. miR-181a, miR-181b, miR-20b, miR-103, miR-15b, miR-331-3p, and let-7c. B. miR-155. C. miR-342-3p, miR-337-3p, miR-198, and miR-34a. For (A, B, C), B (-): control B cell samples. B (+): corresponding activated B samples by CpG. Error bars indicate the standard deviation of 3 independent samples. D, E. miRNA-specific RT-PCR confirms the differential expression of subsets of GSEA-identified miRNAs in CLL relative to control B cells. Upregulated miRNAs are miR-34a, miR-155, miR-198 and miR-342-3p. Downregulated miRNA expression including let-7c, miR-15b, miR-20b, miR-103, miR-181a, miR-181b, miR-331-3p and miR-337-3p. Between 4 to 8 control unstimulated B cell samples (square) and between 6 to 21 CLL patient samples (triangle) were used. The bar indicates the average expression in each group. p-value for the analysis calculated by t-test is shown. miRNA expression was normalized by snoRNA-RNU44 expression. Relative miRNA expression was calculated by ΔΔCt method and is indicated on the Y-axis.

Mentions: Next, we used miRNA-specific RT-PCR to confirm the expression of these signature miRNAs in 3 control B samples and 3 activated B samples (different from the samples used in the miRNA profiling) after CpG activation (Figure 2A, 2B, and 2C) and in control B samples and at least 4 CLL patient samples per miRNA, (Figure 2D and 2E). Using RT-PCR, we independently confirmed these miRNA expression patterns in control B and activated B cells (Figure 2A and 2B). However, in CLL cells, miR-337-3p demonstrated an opposite trend to the one identified in GSEA of miRNA expression profiling (Figure 1E). Moreover, miR-15b and miR-198 demonstrated a similar trend in as in GSEA of miRNA expression profiling (Figure 1C and 1E) though the p value was not statistically significant (Figure 2D and 2E). Variations detected between Luminex bead-based profiling and RT-PCR may be due to the heterogeneity of CLL as we used a subset of CLL samples for the qPCR confirmation.


MicroRNA expression profiling identifies activated B cell status in chronic lymphocytic leukemia cells.

Li S, Moffett HF, Lu J, Werner L, Zhang H, Ritz J, Neuberg D, Wucherpfennig KW, Brown JR, Novina CD - PLoS ONE (2011)

Validation of altered miRNA expression identified by GSEA.miRNA-specific RT-PCR independently confirms altered expression of GSEA-identified miRNAs in B cells activated with CpG. A. miR-181a, miR-181b, miR-20b, miR-103, miR-15b, miR-331-3p, and let-7c. B. miR-155. C. miR-342-3p, miR-337-3p, miR-198, and miR-34a. For (A, B, C), B (-): control B cell samples. B (+): corresponding activated B samples by CpG. Error bars indicate the standard deviation of 3 independent samples. D, E. miRNA-specific RT-PCR confirms the differential expression of subsets of GSEA-identified miRNAs in CLL relative to control B cells. Upregulated miRNAs are miR-34a, miR-155, miR-198 and miR-342-3p. Downregulated miRNA expression including let-7c, miR-15b, miR-20b, miR-103, miR-181a, miR-181b, miR-331-3p and miR-337-3p. Between 4 to 8 control unstimulated B cell samples (square) and between 6 to 21 CLL patient samples (triangle) were used. The bar indicates the average expression in each group. p-value for the analysis calculated by t-test is shown. miRNA expression was normalized by snoRNA-RNU44 expression. Relative miRNA expression was calculated by ΔΔCt method and is indicated on the Y-axis.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3050979&req=5

pone-0016956-g002: Validation of altered miRNA expression identified by GSEA.miRNA-specific RT-PCR independently confirms altered expression of GSEA-identified miRNAs in B cells activated with CpG. A. miR-181a, miR-181b, miR-20b, miR-103, miR-15b, miR-331-3p, and let-7c. B. miR-155. C. miR-342-3p, miR-337-3p, miR-198, and miR-34a. For (A, B, C), B (-): control B cell samples. B (+): corresponding activated B samples by CpG. Error bars indicate the standard deviation of 3 independent samples. D, E. miRNA-specific RT-PCR confirms the differential expression of subsets of GSEA-identified miRNAs in CLL relative to control B cells. Upregulated miRNAs are miR-34a, miR-155, miR-198 and miR-342-3p. Downregulated miRNA expression including let-7c, miR-15b, miR-20b, miR-103, miR-181a, miR-181b, miR-331-3p and miR-337-3p. Between 4 to 8 control unstimulated B cell samples (square) and between 6 to 21 CLL patient samples (triangle) were used. The bar indicates the average expression in each group. p-value for the analysis calculated by t-test is shown. miRNA expression was normalized by snoRNA-RNU44 expression. Relative miRNA expression was calculated by ΔΔCt method and is indicated on the Y-axis.
Mentions: Next, we used miRNA-specific RT-PCR to confirm the expression of these signature miRNAs in 3 control B samples and 3 activated B samples (different from the samples used in the miRNA profiling) after CpG activation (Figure 2A, 2B, and 2C) and in control B samples and at least 4 CLL patient samples per miRNA, (Figure 2D and 2E). Using RT-PCR, we independently confirmed these miRNA expression patterns in control B and activated B cells (Figure 2A and 2B). However, in CLL cells, miR-337-3p demonstrated an opposite trend to the one identified in GSEA of miRNA expression profiling (Figure 1E). Moreover, miR-15b and miR-198 demonstrated a similar trend in as in GSEA of miRNA expression profiling (Figure 1C and 1E) though the p value was not statistically significant (Figure 2D and 2E). Variations detected between Luminex bead-based profiling and RT-PCR may be due to the heterogeneity of CLL as we used a subset of CLL samples for the qPCR confirmation.

Bottom Line: Gene set enrichment analysis (GSEA) identified statistically-significant similarities in miRNA expression between activated B cells and CLL cells including upregulation of miR-34a, miR-155, and miR-342-3p and downregulation of miR-103, miR-181a and miR-181b.Additionally, decreased levels of two CLL signature miRNAs miR-29c and miR-223 are associated with ZAP70(+) and IgV(H) unmutated status and with shorter time to first therapy.These data indicate an activated B cell status for CLL cells and suggest that the direction of change of individual miRNAs may predict clinical course in CLL.

View Article: PubMed Central - PubMed

Affiliation: Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Boston, Massachusetts, United States of America.

ABSTRACT
Chronic lymphocytic leukemia (CLL) is thought to be a disease of resting lymphocytes. However, recent data suggest that CLL cells may more closely resemble activated B cells. Using microRNA (miRNA) expression profiling of highly-enriched CLL cells from 38 patients and 9 untransformed B cells from normal donors before acute CpG activation and 5 matched B cells after acute CpG activation, we demonstrate an activated B cell status for CLL. Gene set enrichment analysis (GSEA) identified statistically-significant similarities in miRNA expression between activated B cells and CLL cells including upregulation of miR-34a, miR-155, and miR-342-3p and downregulation of miR-103, miR-181a and miR-181b. Additionally, decreased levels of two CLL signature miRNAs miR-29c and miR-223 are associated with ZAP70(+) and IgV(H) unmutated status and with shorter time to first therapy. These data indicate an activated B cell status for CLL cells and suggest that the direction of change of individual miRNAs may predict clinical course in CLL.

Show MeSH
Related in: MedlinePlus