Limits...
Hypoxia induces a lipogenic cancer cell phenotype via HIF1α-dependent and -independent pathways.

Valli A, Rodriguez M, Moutsianas L, Fischer R, Fedele V, Huang HL, Van Stiphout R, Jones D, Mccarthy M, Vinaxia M, Igarashi K, Sato M, Soga T, Buffa F, Mccullagh J, Yanes O, Harris A, Kessler B - Oncotarget (2015)

Bottom Line: To study the role of HIF1α in these processes, we used HCT116 colorectal cancer cells expressing endogenous HIF1α and cells in which the hif1α gene was deleted to characterize HIF1α-dependent and independent effects on hypoxia regulated lipid metabolites.Palmitate, stearate, PLD3 and PAFC16 were regulated in a HIF-independent manner.Our results demonstrate the impact of hypoxia on lipid metabolites, of which a distinct subset is regulated by HIF1α.

View Article: PubMed Central - PubMed

Affiliation: Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK.

ABSTRACT
The biochemistry of cancer cells diverges significantly from normal cells as a result of a comprehensive reprogramming of metabolic pathways. A major factor influencing cancer metabolism is hypoxia, which is mediated by HIF1α and HIF2α. HIF1α represents one of the principal regulators of metabolism and energetic balance in cancer cells through its regulation of glycolysis, glycogen synthesis, Krebs cycle and the pentose phosphate shunt. However, less is known about the role of HIF1α in modulating lipid metabolism. Lipids serve cancer cells to provide molecules acting as oncogenic signals, energetic reserve, precursors for new membrane synthesis and to balance redox biological reactions. To study the role of HIF1α in these processes, we used HCT116 colorectal cancer cells expressing endogenous HIF1α and cells in which the hif1α gene was deleted to characterize HIF1α-dependent and independent effects on hypoxia regulated lipid metabolites. Untargeted metabolomics integrated with proteomics revealed that hypoxia induced many changes in lipids metabolites. Enzymatic steps in fatty acid synthesis and the Kennedy pathway were modified in a HIF1α-dependent fashion. Palmitate, stearate, PLD3 and PAFC16 were regulated in a HIF-independent manner. Our results demonstrate the impact of hypoxia on lipid metabolites, of which a distinct subset is regulated by HIF1α.

Show MeSH

Related in: MedlinePlus

Hypoxia-dependent lipophilic molecular features phenotype of HCT116 colorectal cancer cells(a) Biological triplicates of logarithmically growing HCT116 colorectal cancer cells were plated in equal numbers in 21% O2 and 1% O2 and collected after 24 hours with a confluence below 85%. Cell numbers are given as a percentage ±sd relative to the number of HIF1α wild type cells in normoxia, set as 100%. (b) HCT116 cells were treated as described above, HIF1α and HIF2α levels detected by western blot analysis induction in hypoxic. HIF1α increased about eight-fold in wild type hypoxic cells compared to normoxia, and no signal was observed in hif1α−/− cells. HIF2α did not show any compensatory induction in the hif1α−/− cells. (c) In addition, the percentage ±sd of treated cells in G0/G1, S and G2/M phase of the cell cycle is indicated. Representative DNA profiles are presented. HCT116 cells wild type and hif1α−/−, normoxic and hypoxic cells were cytofluorometrically investigated for the percentage ±sd of apoptotic cells with sub-G1 DNA content. (d) Heatmap plotting log2 of average relative intensities of 1,487 detected and baseline normalized untargeted molecular features selected with a fold change ≥2 (in at least one group) and a p<0.05. Untargeted analysis was conducted by LC/MS QTOF nanoflow on the molecular features in the lipophilic phase with positive mode [+] acquisition. (e) Three dimension Principal Component Analysis plotting the data matrix obtained from the selected 1,487 untargeted molecular features. PCA analysis revealed a differential regulation of molecular features in 1% O2 and dependency on the presence of HIF1α (n=5). Percentage of coverage of three principal components analysis is reported. Quality controls were obtained by pooling equal amounts of the analyzed samples, were injected during the analysis at fixed intervals, formed an intermediate cluster assessing the repeatability within the analysis (n=5).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4385826&req=5

Figure 1: Hypoxia-dependent lipophilic molecular features phenotype of HCT116 colorectal cancer cells(a) Biological triplicates of logarithmically growing HCT116 colorectal cancer cells were plated in equal numbers in 21% O2 and 1% O2 and collected after 24 hours with a confluence below 85%. Cell numbers are given as a percentage ±sd relative to the number of HIF1α wild type cells in normoxia, set as 100%. (b) HCT116 cells were treated as described above, HIF1α and HIF2α levels detected by western blot analysis induction in hypoxic. HIF1α increased about eight-fold in wild type hypoxic cells compared to normoxia, and no signal was observed in hif1α−/− cells. HIF2α did not show any compensatory induction in the hif1α−/− cells. (c) In addition, the percentage ±sd of treated cells in G0/G1, S and G2/M phase of the cell cycle is indicated. Representative DNA profiles are presented. HCT116 cells wild type and hif1α−/−, normoxic and hypoxic cells were cytofluorometrically investigated for the percentage ±sd of apoptotic cells with sub-G1 DNA content. (d) Heatmap plotting log2 of average relative intensities of 1,487 detected and baseline normalized untargeted molecular features selected with a fold change ≥2 (in at least one group) and a p<0.05. Untargeted analysis was conducted by LC/MS QTOF nanoflow on the molecular features in the lipophilic phase with positive mode [+] acquisition. (e) Three dimension Principal Component Analysis plotting the data matrix obtained from the selected 1,487 untargeted molecular features. PCA analysis revealed a differential regulation of molecular features in 1% O2 and dependency on the presence of HIF1α (n=5). Percentage of coverage of three principal components analysis is reported. Quality controls were obtained by pooling equal amounts of the analyzed samples, were injected during the analysis at fixed intervals, formed an intermediate cluster assessing the repeatability within the analysis (n=5).

Mentions: Oxygen tension in solid tumors varies considerably between 0.1–2%. In order to reflect this, we chose 1% as the oxygen concentration in our study. Cell proliferation, given as a percentage ±sd relative to the number of HCT116 HIF1α wild type cells in normoxia, was set as 100%. There was a 25%±6% (p-value<0.05) reduction of proliferation observed for HIF1α knockout HCT116 cells (hif1α−/−) at normoxic levels (Figure 1a). Under hypoxic conditions, proliferation of both wild type cells hif1α−/−cells was reduced by 41%±6% (p-value < 0.05) and 47%±11% (p-value<0.01), respectively.


Hypoxia induces a lipogenic cancer cell phenotype via HIF1α-dependent and -independent pathways.

Valli A, Rodriguez M, Moutsianas L, Fischer R, Fedele V, Huang HL, Van Stiphout R, Jones D, Mccarthy M, Vinaxia M, Igarashi K, Sato M, Soga T, Buffa F, Mccullagh J, Yanes O, Harris A, Kessler B - Oncotarget (2015)

Hypoxia-dependent lipophilic molecular features phenotype of HCT116 colorectal cancer cells(a) Biological triplicates of logarithmically growing HCT116 colorectal cancer cells were plated in equal numbers in 21% O2 and 1% O2 and collected after 24 hours with a confluence below 85%. Cell numbers are given as a percentage ±sd relative to the number of HIF1α wild type cells in normoxia, set as 100%. (b) HCT116 cells were treated as described above, HIF1α and HIF2α levels detected by western blot analysis induction in hypoxic. HIF1α increased about eight-fold in wild type hypoxic cells compared to normoxia, and no signal was observed in hif1α−/− cells. HIF2α did not show any compensatory induction in the hif1α−/− cells. (c) In addition, the percentage ±sd of treated cells in G0/G1, S and G2/M phase of the cell cycle is indicated. Representative DNA profiles are presented. HCT116 cells wild type and hif1α−/−, normoxic and hypoxic cells were cytofluorometrically investigated for the percentage ±sd of apoptotic cells with sub-G1 DNA content. (d) Heatmap plotting log2 of average relative intensities of 1,487 detected and baseline normalized untargeted molecular features selected with a fold change ≥2 (in at least one group) and a p<0.05. Untargeted analysis was conducted by LC/MS QTOF nanoflow on the molecular features in the lipophilic phase with positive mode [+] acquisition. (e) Three dimension Principal Component Analysis plotting the data matrix obtained from the selected 1,487 untargeted molecular features. PCA analysis revealed a differential regulation of molecular features in 1% O2 and dependency on the presence of HIF1α (n=5). Percentage of coverage of three principal components analysis is reported. Quality controls were obtained by pooling equal amounts of the analyzed samples, were injected during the analysis at fixed intervals, formed an intermediate cluster assessing the repeatability within the analysis (n=5).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4385826&req=5

Figure 1: Hypoxia-dependent lipophilic molecular features phenotype of HCT116 colorectal cancer cells(a) Biological triplicates of logarithmically growing HCT116 colorectal cancer cells were plated in equal numbers in 21% O2 and 1% O2 and collected after 24 hours with a confluence below 85%. Cell numbers are given as a percentage ±sd relative to the number of HIF1α wild type cells in normoxia, set as 100%. (b) HCT116 cells were treated as described above, HIF1α and HIF2α levels detected by western blot analysis induction in hypoxic. HIF1α increased about eight-fold in wild type hypoxic cells compared to normoxia, and no signal was observed in hif1α−/− cells. HIF2α did not show any compensatory induction in the hif1α−/− cells. (c) In addition, the percentage ±sd of treated cells in G0/G1, S and G2/M phase of the cell cycle is indicated. Representative DNA profiles are presented. HCT116 cells wild type and hif1α−/−, normoxic and hypoxic cells were cytofluorometrically investigated for the percentage ±sd of apoptotic cells with sub-G1 DNA content. (d) Heatmap plotting log2 of average relative intensities of 1,487 detected and baseline normalized untargeted molecular features selected with a fold change ≥2 (in at least one group) and a p<0.05. Untargeted analysis was conducted by LC/MS QTOF nanoflow on the molecular features in the lipophilic phase with positive mode [+] acquisition. (e) Three dimension Principal Component Analysis plotting the data matrix obtained from the selected 1,487 untargeted molecular features. PCA analysis revealed a differential regulation of molecular features in 1% O2 and dependency on the presence of HIF1α (n=5). Percentage of coverage of three principal components analysis is reported. Quality controls were obtained by pooling equal amounts of the analyzed samples, were injected during the analysis at fixed intervals, formed an intermediate cluster assessing the repeatability within the analysis (n=5).
Mentions: Oxygen tension in solid tumors varies considerably between 0.1–2%. In order to reflect this, we chose 1% as the oxygen concentration in our study. Cell proliferation, given as a percentage ±sd relative to the number of HCT116 HIF1α wild type cells in normoxia, was set as 100%. There was a 25%±6% (p-value<0.05) reduction of proliferation observed for HIF1α knockout HCT116 cells (hif1α−/−) at normoxic levels (Figure 1a). Under hypoxic conditions, proliferation of both wild type cells hif1α−/−cells was reduced by 41%±6% (p-value < 0.05) and 47%±11% (p-value<0.01), respectively.

Bottom Line: To study the role of HIF1α in these processes, we used HCT116 colorectal cancer cells expressing endogenous HIF1α and cells in which the hif1α gene was deleted to characterize HIF1α-dependent and independent effects on hypoxia regulated lipid metabolites.Palmitate, stearate, PLD3 and PAFC16 were regulated in a HIF-independent manner.Our results demonstrate the impact of hypoxia on lipid metabolites, of which a distinct subset is regulated by HIF1α.

View Article: PubMed Central - PubMed

Affiliation: Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK.

ABSTRACT
The biochemistry of cancer cells diverges significantly from normal cells as a result of a comprehensive reprogramming of metabolic pathways. A major factor influencing cancer metabolism is hypoxia, which is mediated by HIF1α and HIF2α. HIF1α represents one of the principal regulators of metabolism and energetic balance in cancer cells through its regulation of glycolysis, glycogen synthesis, Krebs cycle and the pentose phosphate shunt. However, less is known about the role of HIF1α in modulating lipid metabolism. Lipids serve cancer cells to provide molecules acting as oncogenic signals, energetic reserve, precursors for new membrane synthesis and to balance redox biological reactions. To study the role of HIF1α in these processes, we used HCT116 colorectal cancer cells expressing endogenous HIF1α and cells in which the hif1α gene was deleted to characterize HIF1α-dependent and independent effects on hypoxia regulated lipid metabolites. Untargeted metabolomics integrated with proteomics revealed that hypoxia induced many changes in lipids metabolites. Enzymatic steps in fatty acid synthesis and the Kennedy pathway were modified in a HIF1α-dependent fashion. Palmitate, stearate, PLD3 and PAFC16 were regulated in a HIF-independent manner. Our results demonstrate the impact of hypoxia on lipid metabolites, of which a distinct subset is regulated by HIF1α.

Show MeSH
Related in: MedlinePlus