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Paradoxical regulation of hypoxia inducible factor-1α (HIF-1α) by histone deacetylase inhibitor in diffuse large B-cell lymphoma.

Bhalla S, Evens AM, Prachand S, Schumacker PT, Gordon LI - PLoS ONE (2013)

Bottom Line: We show here that HIF-1α expression is constitutively active in multiple diffuse large B cell lymphoma (DLBCL) cell lines under normoxia and it is regulated by the PI3K/AKT pathway.We tested the hypothesis that PCI-24781- induced autophagy is mediated by HIF-1α and that inhibition of HIF-1α in these cells results in attenuation of autophagy and decreased survival.We also provide evidence that autophagy serves as a survival pathway in DLBCL cells treated with PCI-24781 which suggests that the use of autophagy inhibitors such as chloroquine or 3-methyl adenine in combination with PCI-24781 may enhance apoptosis in lymphoma cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Division of Hematology/Oncology Northwestern University Feinberg School of Medicine and the Robert H Lurie Comprehensive Cancer Center, Chicago, Illinois, United States of America.

ABSTRACT
Hypoxia inducible factor (HIF) is important in cancer, as it regulates various oncogenic genes as well as genes involved in cell survival, proliferation, and migration. Elevated HIF-1 protein promotes a more aggressive tumor phenotype, and greater HIF-1 expression has been demonstrated to correlate with poorer prognosis, increased risk of metastasis and increased mortality. Recent reports suggest that HIF-1 activates autophagy, a lysosomal degradation pathway which may promote tumor cell survival. We show here that HIF-1α expression is constitutively active in multiple diffuse large B cell lymphoma (DLBCL) cell lines under normoxia and it is regulated by the PI3K/AKT pathway. PCI-24781, a pan histone deacetylase inhibitor (HDACI), enhanced accumulation of HIF-1α and induced autophagy initially, while extended incubation with the drug resulted in inhibition of HIF-1α. We tested the hypothesis that PCI-24781- induced autophagy is mediated by HIF-1α and that inhibition of HIF-1α in these cells results in attenuation of autophagy and decreased survival. We also provide evidence that autophagy serves as a survival pathway in DLBCL cells treated with PCI-24781 which suggests that the use of autophagy inhibitors such as chloroquine or 3-methyl adenine in combination with PCI-24781 may enhance apoptosis in lymphoma cells.

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PCI-24781- Induced autophagy is mediated by HIF-1α.(A) SUDH4, SUDHL6 and OCI-LY3 cells were incubated with increasing concentration of PCI-24781 for 6 or 16 hr followed by cell lysis and western blotting for LC3B and p62 protein using specific antibodies. (B) SUDHL4 and OCI-LY3 cells were incubated with 0.5 or 1.0 µM of PCI respectively for 0-16 hr followed by cell lysis and western blotting for LC3B protein using specific antibody. (C) SUDHL4 and OCI-LY3 cells expressing scrambled or HIF-1α shRNA were treated with PCI for 24 hr followed by further incubation with acridine orange for 15 minute. Cells were washed twice with PBS and analyzed by flow cytomerty for the quantification of cells with high AVO population. (D) OCI-LY3 cells with or without HIF-1α shRNA were treated with PCI-24781 for 6 hr followed by cell lysis and western blotting for autophagy markers p62 and LC3B proteins. Actin is used as an internal control for western blotting. (* = P<0.05; **= p<0.01; *** = P<0.001).
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pone-0081333-g007: PCI-24781- Induced autophagy is mediated by HIF-1α.(A) SUDH4, SUDHL6 and OCI-LY3 cells were incubated with increasing concentration of PCI-24781 for 6 or 16 hr followed by cell lysis and western blotting for LC3B and p62 protein using specific antibodies. (B) SUDHL4 and OCI-LY3 cells were incubated with 0.5 or 1.0 µM of PCI respectively for 0-16 hr followed by cell lysis and western blotting for LC3B protein using specific antibody. (C) SUDHL4 and OCI-LY3 cells expressing scrambled or HIF-1α shRNA were treated with PCI for 24 hr followed by further incubation with acridine orange for 15 minute. Cells were washed twice with PBS and analyzed by flow cytomerty for the quantification of cells with high AVO population. (D) OCI-LY3 cells with or without HIF-1α shRNA were treated with PCI-24781 for 6 hr followed by cell lysis and western blotting for autophagy markers p62 and LC3B proteins. Actin is used as an internal control for western blotting. (* = P<0.05; **= p<0.01; *** = P<0.001).

Mentions: Finally, we examined whether HIF-1α, an upstream activator of autophagy, was involved in PCI-24781-induced autophagy [41]. We initially demonstrated that PCI-24781 caused an upregulation of HIF-1α expression within 6 hr. while extended incubation for 16 hr. with the drug resulted in HIF-1α downregulation. As shown in Figure 7A, a downregulation of the pro-autophagy protein p62 and an increase in LC3B cleavage was also observed at 6hr suggesting that induction of autophagy coincides with increases in HIF-1α expression. Figure 7B shows an increase in LC3BII expression from 0-6hr after treatment with PCI-24781 in both SUDH4 and OCI-LY3 cells.


Paradoxical regulation of hypoxia inducible factor-1α (HIF-1α) by histone deacetylase inhibitor in diffuse large B-cell lymphoma.

Bhalla S, Evens AM, Prachand S, Schumacker PT, Gordon LI - PLoS ONE (2013)

PCI-24781- Induced autophagy is mediated by HIF-1α.(A) SUDH4, SUDHL6 and OCI-LY3 cells were incubated with increasing concentration of PCI-24781 for 6 or 16 hr followed by cell lysis and western blotting for LC3B and p62 protein using specific antibodies. (B) SUDHL4 and OCI-LY3 cells were incubated with 0.5 or 1.0 µM of PCI respectively for 0-16 hr followed by cell lysis and western blotting for LC3B protein using specific antibody. (C) SUDHL4 and OCI-LY3 cells expressing scrambled or HIF-1α shRNA were treated with PCI for 24 hr followed by further incubation with acridine orange for 15 minute. Cells were washed twice with PBS and analyzed by flow cytomerty for the quantification of cells with high AVO population. (D) OCI-LY3 cells with or without HIF-1α shRNA were treated with PCI-24781 for 6 hr followed by cell lysis and western blotting for autophagy markers p62 and LC3B proteins. Actin is used as an internal control for western blotting. (* = P<0.05; **= p<0.01; *** = P<0.001).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3842257&req=5

pone-0081333-g007: PCI-24781- Induced autophagy is mediated by HIF-1α.(A) SUDH4, SUDHL6 and OCI-LY3 cells were incubated with increasing concentration of PCI-24781 for 6 or 16 hr followed by cell lysis and western blotting for LC3B and p62 protein using specific antibodies. (B) SUDHL4 and OCI-LY3 cells were incubated with 0.5 or 1.0 µM of PCI respectively for 0-16 hr followed by cell lysis and western blotting for LC3B protein using specific antibody. (C) SUDHL4 and OCI-LY3 cells expressing scrambled or HIF-1α shRNA were treated with PCI for 24 hr followed by further incubation with acridine orange for 15 minute. Cells were washed twice with PBS and analyzed by flow cytomerty for the quantification of cells with high AVO population. (D) OCI-LY3 cells with or without HIF-1α shRNA were treated with PCI-24781 for 6 hr followed by cell lysis and western blotting for autophagy markers p62 and LC3B proteins. Actin is used as an internal control for western blotting. (* = P<0.05; **= p<0.01; *** = P<0.001).
Mentions: Finally, we examined whether HIF-1α, an upstream activator of autophagy, was involved in PCI-24781-induced autophagy [41]. We initially demonstrated that PCI-24781 caused an upregulation of HIF-1α expression within 6 hr. while extended incubation for 16 hr. with the drug resulted in HIF-1α downregulation. As shown in Figure 7A, a downregulation of the pro-autophagy protein p62 and an increase in LC3B cleavage was also observed at 6hr suggesting that induction of autophagy coincides with increases in HIF-1α expression. Figure 7B shows an increase in LC3BII expression from 0-6hr after treatment with PCI-24781 in both SUDH4 and OCI-LY3 cells.

Bottom Line: We show here that HIF-1α expression is constitutively active in multiple diffuse large B cell lymphoma (DLBCL) cell lines under normoxia and it is regulated by the PI3K/AKT pathway.We tested the hypothesis that PCI-24781- induced autophagy is mediated by HIF-1α and that inhibition of HIF-1α in these cells results in attenuation of autophagy and decreased survival.We also provide evidence that autophagy serves as a survival pathway in DLBCL cells treated with PCI-24781 which suggests that the use of autophagy inhibitors such as chloroquine or 3-methyl adenine in combination with PCI-24781 may enhance apoptosis in lymphoma cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Division of Hematology/Oncology Northwestern University Feinberg School of Medicine and the Robert H Lurie Comprehensive Cancer Center, Chicago, Illinois, United States of America.

ABSTRACT
Hypoxia inducible factor (HIF) is important in cancer, as it regulates various oncogenic genes as well as genes involved in cell survival, proliferation, and migration. Elevated HIF-1 protein promotes a more aggressive tumor phenotype, and greater HIF-1 expression has been demonstrated to correlate with poorer prognosis, increased risk of metastasis and increased mortality. Recent reports suggest that HIF-1 activates autophagy, a lysosomal degradation pathway which may promote tumor cell survival. We show here that HIF-1α expression is constitutively active in multiple diffuse large B cell lymphoma (DLBCL) cell lines under normoxia and it is regulated by the PI3K/AKT pathway. PCI-24781, a pan histone deacetylase inhibitor (HDACI), enhanced accumulation of HIF-1α and induced autophagy initially, while extended incubation with the drug resulted in inhibition of HIF-1α. We tested the hypothesis that PCI-24781- induced autophagy is mediated by HIF-1α and that inhibition of HIF-1α in these cells results in attenuation of autophagy and decreased survival. We also provide evidence that autophagy serves as a survival pathway in DLBCL cells treated with PCI-24781 which suggests that the use of autophagy inhibitors such as chloroquine or 3-methyl adenine in combination with PCI-24781 may enhance apoptosis in lymphoma cells.

Show MeSH
Related in: MedlinePlus