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The ATRA-induced differentiation of medulloblastoma cells is enhanced with LOX/COX inhibitors: an analysis of gene expression.

Chlapek P, Neradil J, Redova M, Zitterbart K, Sterba J, Veselska R - Cancer Cell Int. (2014)

Bottom Line: This effect was detected in both cell lines.This effect was also observed for the CDKN1A gene encoding the p21 protein, which is an important regulator of the cell cycle, and for the genes encoding proteins that are associated with proteasome activity.Furthermore, our results showed that D283 Med cells are significantly more sensitive to treatment with ATRA alone than Daoy cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Experimental Biology - Laboratory of Tumor Biology, School of Science, Masaryk University, Kotlarska 2, 611 37 Brno, Czech Republic.

ABSTRACT

Background: A detailed analysis of the expression of 440 cancer-related genes was performed after the combined treatment of medulloblastoma cells with all-trans retinoic acid (ATRA) and inhibitors of lipoxygenases (LOX) and cyclooxygenases (COX). The combinations of retinoids and celecoxib as a COX-2 inhibitor were reported to be effective in some regimens of metronomic therapy of relapsed solid tumors with poor prognosis. Our previous findings on neuroblastoma cells using expression profiling showed that LOX/COX inhibitors have the capability of enhancing the differentiating action of ATRA. Presented study focused on the continuation of our previous work to confirm the possibility of enhancing ATRA-induced cell differentiation in these cell lines via the application of LOX/COX inhibitors. This study provides more detailed information concerning the mechanisms of the enhancement of the ATRA-induced differentiation of medulloblastoma cells.

Methods: The Daoy and D283 Med medulloblastoma cell lines were chosen for this study. Caffeic acid (an inhibitor of 5-LOX) and celecoxib (an inhibitor on COX-2) were used in combined treatment with ATRA. The expression profiling was performed using Human Cancer Oligo GEArray membranes, and the most promising results were verified using RT-PCR.

Results: The expression profiling of the selected cancer-related genes clearly confirmed that the differentiating effects of ATRA should be enhanced via its combined administration with caffeic acid or celecoxib. This effect was detected in both cell lines. An increased expression of the genes that encoded the proteins participating in induced differentiation and cytoskeleton remodeling was detected in both cell lines in a concentration-dependent manner. This effect was also observed for the CDKN1A gene encoding the p21 protein, which is an important regulator of the cell cycle, and for the genes encoding proteins that are associated with proteasome activity. Furthermore, our results showed that D283 Med cells are significantly more sensitive to treatment with ATRA alone than Daoy cells.

Conclusions: The obtained results on medulloblastoma cell lines are in accordance with our previous findings on neuroblastoma cells and confirm our hypothesis concerning the common mechanism of the enhancement of ATRA-induced cell differentiation in various types of pediatric solid tumors.

No MeSH data available.


Related in: MedlinePlus

Results of gene cluster analysis after the expression profiling of treated cells. The genes were clustered according to the type of changes in the expression in the respective cell line (Daoy or D283 Med) after the combined treatment with ATRA and inhibitors (CA or CX). The cells were treated with ATRA alone or in combination with CA (an inhibitor of 5-LOX) or CX (an inhibitor of COX-2); numbers indicate the concentration in μM. The green color at the farthest left end of the color scale corresponds to the minimal value; the red color at the farthest right end of the color scale corresponds to the maximal value; the black color in the middle corresponds to the average value. Each of the other values corresponds to a certain color according to its magnitude. The colors are assigned according to the value of the particular gene expression in all samples in the respective experimental variant (I, II, III or IV). Genes discussed in the text in detail are highlighted by red dots.
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Figure 2: Results of gene cluster analysis after the expression profiling of treated cells. The genes were clustered according to the type of changes in the expression in the respective cell line (Daoy or D283 Med) after the combined treatment with ATRA and inhibitors (CA or CX). The cells were treated with ATRA alone or in combination with CA (an inhibitor of 5-LOX) or CX (an inhibitor of COX-2); numbers indicate the concentration in μM. The green color at the farthest left end of the color scale corresponds to the minimal value; the red color at the farthest right end of the color scale corresponds to the maximal value; the black color in the middle corresponds to the average value. Each of the other values corresponds to a certain color according to its magnitude. The colors are assigned according to the value of the particular gene expression in all samples in the respective experimental variant (I, II, III or IV). Genes discussed in the text in detail are highlighted by red dots.

Mentions: The present study was focused on a detailed analysis of Daoy and D283 Med medulloblastoma cells after the combined application of ATRA and LOX/COX inhibitors. CA as the specific inhibitor of 5-LOX and CX as the specific inhibitor of COX-2 were used in these experiments. The changes in the expression of cancer-related genes were evaluated using expression profiling. Furthermore, a detailed analysis of the expression of five candidate genes was performed using RT-PCR to verify the microarray results. We used the same experimental design as our previous studies on neuroblastoma cells [20,29].In Daoy cells, changes in the expression of 80 cancer-related genes were detected after combined treatment with ATRA and inhibitors (Figure 1A). A total of 29 of these genes demonstrated changed expressions after combinations of ATRA and CA as well as of ATRA and CX. The expressions of another 29 genes were changed only after the combined treatment of ATRA and CA. A total of 22 different genes showed changes in expression after undergoing combined treatment with ATRA and CX (Figure 1A).In D283 Med cells, the expressions of 37 genes were changed (Figure 1B). Of these, 22 showed changes after combined treatment with ATRA and CA as well as with ATRA and CX. Changes in the expression of another 11 genes were identified after treatment with ATRA and CA only. Similarly, the expressions of 4 different genes were changed after treatment with ATRA and CX only (Figure 1B).The data achieved via expression profiling are presented after cluster analysis, which grouped the genes or gene groups by the type of changes in their expressions (Figure 2). Based on this analysis, three typical patterns of changes in gene expression are described:


The ATRA-induced differentiation of medulloblastoma cells is enhanced with LOX/COX inhibitors: an analysis of gene expression.

Chlapek P, Neradil J, Redova M, Zitterbart K, Sterba J, Veselska R - Cancer Cell Int. (2014)

Results of gene cluster analysis after the expression profiling of treated cells. The genes were clustered according to the type of changes in the expression in the respective cell line (Daoy or D283 Med) after the combined treatment with ATRA and inhibitors (CA or CX). The cells were treated with ATRA alone or in combination with CA (an inhibitor of 5-LOX) or CX (an inhibitor of COX-2); numbers indicate the concentration in μM. The green color at the farthest left end of the color scale corresponds to the minimal value; the red color at the farthest right end of the color scale corresponds to the maximal value; the black color in the middle corresponds to the average value. Each of the other values corresponds to a certain color according to its magnitude. The colors are assigned according to the value of the particular gene expression in all samples in the respective experimental variant (I, II, III or IV). Genes discussed in the text in detail are highlighted by red dots.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4066709&req=5

Figure 2: Results of gene cluster analysis after the expression profiling of treated cells. The genes were clustered according to the type of changes in the expression in the respective cell line (Daoy or D283 Med) after the combined treatment with ATRA and inhibitors (CA or CX). The cells were treated with ATRA alone or in combination with CA (an inhibitor of 5-LOX) or CX (an inhibitor of COX-2); numbers indicate the concentration in μM. The green color at the farthest left end of the color scale corresponds to the minimal value; the red color at the farthest right end of the color scale corresponds to the maximal value; the black color in the middle corresponds to the average value. Each of the other values corresponds to a certain color according to its magnitude. The colors are assigned according to the value of the particular gene expression in all samples in the respective experimental variant (I, II, III or IV). Genes discussed in the text in detail are highlighted by red dots.
Mentions: The present study was focused on a detailed analysis of Daoy and D283 Med medulloblastoma cells after the combined application of ATRA and LOX/COX inhibitors. CA as the specific inhibitor of 5-LOX and CX as the specific inhibitor of COX-2 were used in these experiments. The changes in the expression of cancer-related genes were evaluated using expression profiling. Furthermore, a detailed analysis of the expression of five candidate genes was performed using RT-PCR to verify the microarray results. We used the same experimental design as our previous studies on neuroblastoma cells [20,29].In Daoy cells, changes in the expression of 80 cancer-related genes were detected after combined treatment with ATRA and inhibitors (Figure 1A). A total of 29 of these genes demonstrated changed expressions after combinations of ATRA and CA as well as of ATRA and CX. The expressions of another 29 genes were changed only after the combined treatment of ATRA and CA. A total of 22 different genes showed changes in expression after undergoing combined treatment with ATRA and CX (Figure 1A).In D283 Med cells, the expressions of 37 genes were changed (Figure 1B). Of these, 22 showed changes after combined treatment with ATRA and CA as well as with ATRA and CX. Changes in the expression of another 11 genes were identified after treatment with ATRA and CA only. Similarly, the expressions of 4 different genes were changed after treatment with ATRA and CX only (Figure 1B).The data achieved via expression profiling are presented after cluster analysis, which grouped the genes or gene groups by the type of changes in their expressions (Figure 2). Based on this analysis, three typical patterns of changes in gene expression are described:

Bottom Line: This effect was detected in both cell lines.This effect was also observed for the CDKN1A gene encoding the p21 protein, which is an important regulator of the cell cycle, and for the genes encoding proteins that are associated with proteasome activity.Furthermore, our results showed that D283 Med cells are significantly more sensitive to treatment with ATRA alone than Daoy cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Experimental Biology - Laboratory of Tumor Biology, School of Science, Masaryk University, Kotlarska 2, 611 37 Brno, Czech Republic.

ABSTRACT

Background: A detailed analysis of the expression of 440 cancer-related genes was performed after the combined treatment of medulloblastoma cells with all-trans retinoic acid (ATRA) and inhibitors of lipoxygenases (LOX) and cyclooxygenases (COX). The combinations of retinoids and celecoxib as a COX-2 inhibitor were reported to be effective in some regimens of metronomic therapy of relapsed solid tumors with poor prognosis. Our previous findings on neuroblastoma cells using expression profiling showed that LOX/COX inhibitors have the capability of enhancing the differentiating action of ATRA. Presented study focused on the continuation of our previous work to confirm the possibility of enhancing ATRA-induced cell differentiation in these cell lines via the application of LOX/COX inhibitors. This study provides more detailed information concerning the mechanisms of the enhancement of the ATRA-induced differentiation of medulloblastoma cells.

Methods: The Daoy and D283 Med medulloblastoma cell lines were chosen for this study. Caffeic acid (an inhibitor of 5-LOX) and celecoxib (an inhibitor on COX-2) were used in combined treatment with ATRA. The expression profiling was performed using Human Cancer Oligo GEArray membranes, and the most promising results were verified using RT-PCR.

Results: The expression profiling of the selected cancer-related genes clearly confirmed that the differentiating effects of ATRA should be enhanced via its combined administration with caffeic acid or celecoxib. This effect was detected in both cell lines. An increased expression of the genes that encoded the proteins participating in induced differentiation and cytoskeleton remodeling was detected in both cell lines in a concentration-dependent manner. This effect was also observed for the CDKN1A gene encoding the p21 protein, which is an important regulator of the cell cycle, and for the genes encoding proteins that are associated with proteasome activity. Furthermore, our results showed that D283 Med cells are significantly more sensitive to treatment with ATRA alone than Daoy cells.

Conclusions: The obtained results on medulloblastoma cell lines are in accordance with our previous findings on neuroblastoma cells and confirm our hypothesis concerning the common mechanism of the enhancement of ATRA-induced cell differentiation in various types of pediatric solid tumors.

No MeSH data available.


Related in: MedlinePlus