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EMT-associated factors promote invasive properties of uveal melanoma cells.

Asnaghi L, Gezgin G, Tripathy A, Handa JT, Merbs SL, van der Velden PA, Jager MJ, Harbour JW, Eberhart CG - Mol. Vis. (2015)

Bottom Line: The genetic downregulation of ZEB1 in OCM1, OMM1, and 92.1 resulted in a more than 50% reduction in invasion, but only suppressed growth in OMM1 cells.Suppression of Twist1 in Mel290 and OMM1 reduced growth and invasion by more than 50%.The downregulation of Snail1 in the 92.1 cell line reduced invasion by 50%, but did not interfere with growth.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Johns Hopkins University, School of Medicine, Baltimore, MD.

ABSTRACT

Purpose: Transcription factors regulating the epithelial-to-mesenchymal transition (EMT) program contribute to carcinogenesis and metastasis in many tumors, including cutaneous melanoma. However, little is known about the role of EMT factors in the growth and metastatic dissemination of uveal melanoma cells. Here, we analyzed the expression and functions of the EMT factors ZEB1, Twist1, and Snail1 in uveal melanoma cell lines and primary tumors.

Methods: ZEB1, Twist1, and Snail1 mRNA levels were measured using qPCR in five uveal melanoma cell lines and in 30 primary tumors. Gene expression was used to determine class 1 and class 2 signatures in the primary tumors. Short hairpin RNA was used to downregulate the expressions of the EMT factors; then, growth and transwell invasion assays were performed.

Results: ZEB1, Twist1, and Snail1 were expressed in all five uveal melanoma lines, with ZEB1 having the highest protein levels. ZEB1 mRNA was significantly elevated in highly metastatic class 2 primary tumors for which survival data were not available, whereas a high gene expression of Twist1 was associated with a worse prognosis in a separate tumor cohort analyzed by expression profiling. The genetic downregulation of ZEB1 in OCM1, OMM1, and 92.1 resulted in a more than 50% reduction in invasion, but only suppressed growth in OMM1 cells. Suppression of Twist1 in Mel290 and OMM1 reduced growth and invasion by more than 50%. The downregulation of Snail1 in the 92.1 cell line reduced invasion by 50%, but did not interfere with growth.

Conclusions: The downregulation of ZEB1, Twist1, and Snail1 reduces the invasive properties of uveal melanoma cells, and the elevated mRNA levels of ZEB1 and Twist1 are associated with a more aggressive clinical phenotype in uveal melanoma samples. Therefore, these factors could represent new therapeutic targets in patients with ocular melanoma.

No MeSH data available.


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Working model for the activity of the EMT factors in the regulation of invasion in uveal melanoma cells.
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f6: Working model for the activity of the EMT factors in the regulation of invasion in uveal melanoma cells.

Mentions: The suppression of Snail1 was achieved in 92.1 cells using two separate shRNAs constructs, which reduced expression by approximately 40%–60% at the mRNA and protein levels, as compared to scramble controls (Figure 5A,B). We attempted to suppress the Snail1 expression in OCM1 cells as well, but we were not successful. The reduction of Snail1 in 92.1 cultures did not interfere with cell growth, as found by MTS assay performed for 3, 5, and 7 days (Figure 5C). Nevertheless, we observed that Snail1 shRNAs reduced by approximately 50% the number of 92.1 cells that moved through a Matrigel-coated filter after 24 h of incubation (Figure 5D), indicating that Snail1 can also promote invasion in uveal melanoma cells. Our working model for the activity of the EMT factors in the regulation of invasion in uveal melanoma cells is depicted in Figure 6.


EMT-associated factors promote invasive properties of uveal melanoma cells.

Asnaghi L, Gezgin G, Tripathy A, Handa JT, Merbs SL, van der Velden PA, Jager MJ, Harbour JW, Eberhart CG - Mol. Vis. (2015)

Working model for the activity of the EMT factors in the regulation of invasion in uveal melanoma cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4548792&req=5

f6: Working model for the activity of the EMT factors in the regulation of invasion in uveal melanoma cells.
Mentions: The suppression of Snail1 was achieved in 92.1 cells using two separate shRNAs constructs, which reduced expression by approximately 40%–60% at the mRNA and protein levels, as compared to scramble controls (Figure 5A,B). We attempted to suppress the Snail1 expression in OCM1 cells as well, but we were not successful. The reduction of Snail1 in 92.1 cultures did not interfere with cell growth, as found by MTS assay performed for 3, 5, and 7 days (Figure 5C). Nevertheless, we observed that Snail1 shRNAs reduced by approximately 50% the number of 92.1 cells that moved through a Matrigel-coated filter after 24 h of incubation (Figure 5D), indicating that Snail1 can also promote invasion in uveal melanoma cells. Our working model for the activity of the EMT factors in the regulation of invasion in uveal melanoma cells is depicted in Figure 6.

Bottom Line: The genetic downregulation of ZEB1 in OCM1, OMM1, and 92.1 resulted in a more than 50% reduction in invasion, but only suppressed growth in OMM1 cells.Suppression of Twist1 in Mel290 and OMM1 reduced growth and invasion by more than 50%.The downregulation of Snail1 in the 92.1 cell line reduced invasion by 50%, but did not interfere with growth.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Johns Hopkins University, School of Medicine, Baltimore, MD.

ABSTRACT

Purpose: Transcription factors regulating the epithelial-to-mesenchymal transition (EMT) program contribute to carcinogenesis and metastasis in many tumors, including cutaneous melanoma. However, little is known about the role of EMT factors in the growth and metastatic dissemination of uveal melanoma cells. Here, we analyzed the expression and functions of the EMT factors ZEB1, Twist1, and Snail1 in uveal melanoma cell lines and primary tumors.

Methods: ZEB1, Twist1, and Snail1 mRNA levels were measured using qPCR in five uveal melanoma cell lines and in 30 primary tumors. Gene expression was used to determine class 1 and class 2 signatures in the primary tumors. Short hairpin RNA was used to downregulate the expressions of the EMT factors; then, growth and transwell invasion assays were performed.

Results: ZEB1, Twist1, and Snail1 were expressed in all five uveal melanoma lines, with ZEB1 having the highest protein levels. ZEB1 mRNA was significantly elevated in highly metastatic class 2 primary tumors for which survival data were not available, whereas a high gene expression of Twist1 was associated with a worse prognosis in a separate tumor cohort analyzed by expression profiling. The genetic downregulation of ZEB1 in OCM1, OMM1, and 92.1 resulted in a more than 50% reduction in invasion, but only suppressed growth in OMM1 cells. Suppression of Twist1 in Mel290 and OMM1 reduced growth and invasion by more than 50%. The downregulation of Snail1 in the 92.1 cell line reduced invasion by 50%, but did not interfere with growth.

Conclusions: The downregulation of ZEB1, Twist1, and Snail1 reduces the invasive properties of uveal melanoma cells, and the elevated mRNA levels of ZEB1 and Twist1 are associated with a more aggressive clinical phenotype in uveal melanoma samples. Therefore, these factors could represent new therapeutic targets in patients with ocular melanoma.

No MeSH data available.


Related in: MedlinePlus