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Omega 3 fatty acids chemosensitize multidrug resistant colon cancer cells by down-regulating cholesterol synthesis and altering detergent resistant membranes composition.

Gelsomino G, Corsetto PA, Campia I, Montorfano G, Kopecka J, Castella B, Gazzano E, Ghigo D, Rizzo AM, Riganti C - Mol. Cancer (2013)

Bottom Line: MDR cells, which overexpressed Pgp and MRP1, had a dysregulated cholesterol metabolism, due to the lower expression of ubiquitin E3 ligase Trc8: this produced lower ubiquitination rate of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCoAR), higher cholesterol synthesis, higher cholesterol content in MDR cells.We found that DHA and EPA re-activated Trc8 E3 ligase in MDR cells, restored the ubiquitination rate of HMGCoAR to levels comparable with chemosensitive cells, reduced the cholesterol synthesis and incorporation in DRMs. Omega 3 PUFAs were incorporated in whole lipids as well as in DRMs of MDR cells, and altered the lipid composition of these compartments.They reduced the amount of Pgp and MRP1 contained in DRMs, decreased the transporters activity, restored the antitumor effects of different chemotherapeutic drugs, restored a proper tumor-immune system recognition in response to chemotherapy in MDR cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Oncology, University of Torino, via Santena 5/bis, 10126 Torino, Italy. dario.ghigo@unito.it.

ABSTRACT

Background: The activity of P-glycoprotein (Pgp) and multidrug resistance related protein 1 (MRP1), two membrane transporters involved in multidrug resistance of colon cancer, is increased by high amounts of cholesterol in plasma membrane and detergent resistant membranes (DRMs). It has never been investigated whether omega 3 polyunsatured fatty acids (PUFAs), which modulate cholesterol homeostasis in dyslipidemic syndromes and have chemopreventive effects in colon cancer, may affect the response to chemotherapy in multidrug resistant (MDR) tumors.

Methods: We studied the effect of omega 3 PUFAs docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) in human chemosensitive colon cancer HT29 cells and in their MDR counterpart, HT29-dx cells.

Results: MDR cells, which overexpressed Pgp and MRP1, had a dysregulated cholesterol metabolism, due to the lower expression of ubiquitin E3 ligase Trc8: this produced lower ubiquitination rate of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCoAR), higher cholesterol synthesis, higher cholesterol content in MDR cells. We found that DHA and EPA re-activated Trc8 E3 ligase in MDR cells, restored the ubiquitination rate of HMGCoAR to levels comparable with chemosensitive cells, reduced the cholesterol synthesis and incorporation in DRMs. Omega 3 PUFAs were incorporated in whole lipids as well as in DRMs of MDR cells, and altered the lipid composition of these compartments. They reduced the amount of Pgp and MRP1 contained in DRMs, decreased the transporters activity, restored the antitumor effects of different chemotherapeutic drugs, restored a proper tumor-immune system recognition in response to chemotherapy in MDR cells.

Conclusions: Our work describes a new biochemical effect of omega 3 PUFAs, which can be useful to overcome chemoresistance in MDR colon cancer cells.

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ω3PUFAs decrease activity and expression of HMGCoAR in chemoresistant colon cancer cells. HT29 and HT29-dx cells were incubated for 24 h in the absence (CTRL) or in the presence of 50 μM arachidonic acid (AA), docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA). Simvastatin (1 μM for 24 h, SIM) was chosen as HMGCoAR inhibitor. Cells were lysed and centrifuged to collect the microsomal fraction, on which the following investigations were performed. (A) HMGCoAR enzymatic activity was measured in duplicate (see Methods). Data are presented as means ± SD (n = 3). Versus CTRL HT29: * p < 0.02; versus CTRL HT29-dx: ° p < 0.05. (B) Western blotting experiments were performed using an anti-HMGCoAR antibody; an anti-calreticulin (CRT) antibody was used as a control of equal protein loading. The figure is representative of three experiments with similar results. The band density ratio between HMGCoAR and CRT was expressed as arbitrary units. Versus CTRL HT29: * p < 0.05; versus CTRL HT29-dx: ° p < 0.05.
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Figure 2: ω3PUFAs decrease activity and expression of HMGCoAR in chemoresistant colon cancer cells. HT29 and HT29-dx cells were incubated for 24 h in the absence (CTRL) or in the presence of 50 μM arachidonic acid (AA), docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA). Simvastatin (1 μM for 24 h, SIM) was chosen as HMGCoAR inhibitor. Cells were lysed and centrifuged to collect the microsomal fraction, on which the following investigations were performed. (A) HMGCoAR enzymatic activity was measured in duplicate (see Methods). Data are presented as means ± SD (n = 3). Versus CTRL HT29: * p < 0.02; versus CTRL HT29-dx: ° p < 0.05. (B) Western blotting experiments were performed using an anti-HMGCoAR antibody; an anti-calreticulin (CRT) antibody was used as a control of equal protein loading. The figure is representative of three experiments with similar results. The band density ratio between HMGCoAR and CRT was expressed as arbitrary units. Versus CTRL HT29: * p < 0.05; versus CTRL HT29-dx: ° p < 0.05.

Mentions: The higher synthesis of cholesterol in HT29-dx cells was accompanied by higher enzymatic activity (Figure 2A) and protein expression (Figure 2B) of HMGCoAR, compared to HT29 cells. Whereas AA did not modify these parameters, DHA and EPA significantly lowered the activity (Figure 2A) and expression (Figure 2B) of HMGCoAR in chemoresistant cells. Interestingly, they had no effect in chemosensitive cells.


Omega 3 fatty acids chemosensitize multidrug resistant colon cancer cells by down-regulating cholesterol synthesis and altering detergent resistant membranes composition.

Gelsomino G, Corsetto PA, Campia I, Montorfano G, Kopecka J, Castella B, Gazzano E, Ghigo D, Rizzo AM, Riganti C - Mol. Cancer (2013)

ω3PUFAs decrease activity and expression of HMGCoAR in chemoresistant colon cancer cells. HT29 and HT29-dx cells were incubated for 24 h in the absence (CTRL) or in the presence of 50 μM arachidonic acid (AA), docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA). Simvastatin (1 μM for 24 h, SIM) was chosen as HMGCoAR inhibitor. Cells were lysed and centrifuged to collect the microsomal fraction, on which the following investigations were performed. (A) HMGCoAR enzymatic activity was measured in duplicate (see Methods). Data are presented as means ± SD (n = 3). Versus CTRL HT29: * p < 0.02; versus CTRL HT29-dx: ° p < 0.05. (B) Western blotting experiments were performed using an anti-HMGCoAR antibody; an anti-calreticulin (CRT) antibody was used as a control of equal protein loading. The figure is representative of three experiments with similar results. The band density ratio between HMGCoAR and CRT was expressed as arbitrary units. Versus CTRL HT29: * p < 0.05; versus CTRL HT29-dx: ° p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4225767&req=5

Figure 2: ω3PUFAs decrease activity and expression of HMGCoAR in chemoresistant colon cancer cells. HT29 and HT29-dx cells were incubated for 24 h in the absence (CTRL) or in the presence of 50 μM arachidonic acid (AA), docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA). Simvastatin (1 μM for 24 h, SIM) was chosen as HMGCoAR inhibitor. Cells were lysed and centrifuged to collect the microsomal fraction, on which the following investigations were performed. (A) HMGCoAR enzymatic activity was measured in duplicate (see Methods). Data are presented as means ± SD (n = 3). Versus CTRL HT29: * p < 0.02; versus CTRL HT29-dx: ° p < 0.05. (B) Western blotting experiments were performed using an anti-HMGCoAR antibody; an anti-calreticulin (CRT) antibody was used as a control of equal protein loading. The figure is representative of three experiments with similar results. The band density ratio between HMGCoAR and CRT was expressed as arbitrary units. Versus CTRL HT29: * p < 0.05; versus CTRL HT29-dx: ° p < 0.05.
Mentions: The higher synthesis of cholesterol in HT29-dx cells was accompanied by higher enzymatic activity (Figure 2A) and protein expression (Figure 2B) of HMGCoAR, compared to HT29 cells. Whereas AA did not modify these parameters, DHA and EPA significantly lowered the activity (Figure 2A) and expression (Figure 2B) of HMGCoAR in chemoresistant cells. Interestingly, they had no effect in chemosensitive cells.

Bottom Line: MDR cells, which overexpressed Pgp and MRP1, had a dysregulated cholesterol metabolism, due to the lower expression of ubiquitin E3 ligase Trc8: this produced lower ubiquitination rate of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCoAR), higher cholesterol synthesis, higher cholesterol content in MDR cells.We found that DHA and EPA re-activated Trc8 E3 ligase in MDR cells, restored the ubiquitination rate of HMGCoAR to levels comparable with chemosensitive cells, reduced the cholesterol synthesis and incorporation in DRMs. Omega 3 PUFAs were incorporated in whole lipids as well as in DRMs of MDR cells, and altered the lipid composition of these compartments.They reduced the amount of Pgp and MRP1 contained in DRMs, decreased the transporters activity, restored the antitumor effects of different chemotherapeutic drugs, restored a proper tumor-immune system recognition in response to chemotherapy in MDR cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Oncology, University of Torino, via Santena 5/bis, 10126 Torino, Italy. dario.ghigo@unito.it.

ABSTRACT

Background: The activity of P-glycoprotein (Pgp) and multidrug resistance related protein 1 (MRP1), two membrane transporters involved in multidrug resistance of colon cancer, is increased by high amounts of cholesterol in plasma membrane and detergent resistant membranes (DRMs). It has never been investigated whether omega 3 polyunsatured fatty acids (PUFAs), which modulate cholesterol homeostasis in dyslipidemic syndromes and have chemopreventive effects in colon cancer, may affect the response to chemotherapy in multidrug resistant (MDR) tumors.

Methods: We studied the effect of omega 3 PUFAs docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) in human chemosensitive colon cancer HT29 cells and in their MDR counterpart, HT29-dx cells.

Results: MDR cells, which overexpressed Pgp and MRP1, had a dysregulated cholesterol metabolism, due to the lower expression of ubiquitin E3 ligase Trc8: this produced lower ubiquitination rate of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCoAR), higher cholesterol synthesis, higher cholesterol content in MDR cells. We found that DHA and EPA re-activated Trc8 E3 ligase in MDR cells, restored the ubiquitination rate of HMGCoAR to levels comparable with chemosensitive cells, reduced the cholesterol synthesis and incorporation in DRMs. Omega 3 PUFAs were incorporated in whole lipids as well as in DRMs of MDR cells, and altered the lipid composition of these compartments. They reduced the amount of Pgp and MRP1 contained in DRMs, decreased the transporters activity, restored the antitumor effects of different chemotherapeutic drugs, restored a proper tumor-immune system recognition in response to chemotherapy in MDR cells.

Conclusions: Our work describes a new biochemical effect of omega 3 PUFAs, which can be useful to overcome chemoresistance in MDR colon cancer cells.

Show MeSH
Related in: MedlinePlus