Limits...
Hypoxia-associated p38 mitogen-activated protein kinase-mediated androgen receptor activation and increased HIF-1alpha levels contribute to emergence of an aggressive phenotype in prostate cancer.

Khandrika L, Lieberman R, Koul S, Kumar B, Maroni P, Chandhoke R, Meacham RB, Koul HK - Oncogene (2009)

Bottom Line: Our results demonstrate that hypoxia-reoxygenation resulted in increased survival, higher clonogenicity and enhanced invasiveness of these cells.Moreover, hypoxia-reoxygenation was associated with an increased AR activity independent of androgens as well as increased hypoxia inducible factor (HIF-1alpha) levels and activity.We also observed that the activation of p38 mitogen-activated protein (MAP) kinase pathway was an early response to hypoxia, and inhibition of p38 MAP kinase pathway by variety of approaches abolished hypoxia-reoxygenation induced increased AR activity as well as increased survival, clonogenicity and invasiveness.

View Article: PubMed Central - PubMed

Affiliation: Signal Transduction and Molecular Urology Laboratory-Program in Urosciences, Division of Urology, Department of Surgery, School of Medicine, University of Colorado at Denver and Health Sciences Center, Aurora, CO 80045, USA.

ABSTRACT
Androgen receptor (AR) signaling is involved in the development and progression of prostate cancer. Tumor microvasculature contributes to continual exposure of prostate cancer cells to hypoxia-reoxygenation, however, the role of hypoxia-reoxygenation in prostate cancer progression and modulation of AR signaling is not understood. In this study, we evaluated the effects of hypoxia-reoxygenation in LNCaP cells, a line of hormone responsive human prostate cancer cells. Our results demonstrate that hypoxia-reoxygenation resulted in increased survival, higher clonogenicity and enhanced invasiveness of these cells. Moreover, hypoxia-reoxygenation was associated with an increased AR activity independent of androgens as well as increased hypoxia inducible factor (HIF-1alpha) levels and activity. We also observed that the activation of p38 mitogen-activated protein (MAP) kinase pathway was an early response to hypoxia, and inhibition of p38 MAP kinase pathway by variety of approaches abolished hypoxia-reoxygenation induced increased AR activity as well as increased survival, clonogenicity and invasiveness. These results demonstrate a critical role for hypoxia-induced p38 MAP kinase pathway in androgen-independent AR activation in prostate cancer cells, and suggest that hypoxia-reoxygenation may select for aggressive androgen-independent prostate cancer phenotype.

Show MeSH

Related in: MedlinePlus

Chronic hypoxia- reoxygenation selects for LNCaP cells with aggressive phenotype(a) LNCaP cells were cultured in complete medium and incubated in hypoxia for 12h and re-oxygenated for 12h, for 3 weeks. Equal numbers of surviving cells (hrLNCaP) were seeded in androgen free medium and proliferation was measured by MTT assay over 3 days. Each data point represents mean ± S.D of two individual experiments done in quadruplicate (*** indicates p<0.001, n=8). (b) Equal number of hrLNCaP cells and LNCaP cells were seeded in Boyden chambers in androgen free medium and cells were allowed to invade either in the presence or absence of 10nM DHT. 48h later, invaded cells were stained with Crystal Violet (Left Panel). Representative images from two individual experiments done in duplicate are shown. Invaded cells were quantified as before and each data point represents mean ± S.D (* indicates p<0.05 compared to LNCaP cells in androgen free medium, n=4) (Right Panel). (c) Plausible mechanism of dual control of p38 MAPK to promote aggressive growth of prostate cancer cells in hypoxia. When subjected to hypoxia, AR can be activated in an androgen independent manner via p38 MAPK and HSP27 and can translocate into the nucleus. On the other hand, activated p38 MAPK can help stabilize HIF-1α and promote HIF-1 mediated gene transcription. Together the active AR and HIF-1 promote aggressive growth and may in due course lead to androgen independent prostate cancer.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2651999&req=5

Figure 6: Chronic hypoxia- reoxygenation selects for LNCaP cells with aggressive phenotype(a) LNCaP cells were cultured in complete medium and incubated in hypoxia for 12h and re-oxygenated for 12h, for 3 weeks. Equal numbers of surviving cells (hrLNCaP) were seeded in androgen free medium and proliferation was measured by MTT assay over 3 days. Each data point represents mean ± S.D of two individual experiments done in quadruplicate (*** indicates p<0.001, n=8). (b) Equal number of hrLNCaP cells and LNCaP cells were seeded in Boyden chambers in androgen free medium and cells were allowed to invade either in the presence or absence of 10nM DHT. 48h later, invaded cells were stained with Crystal Violet (Left Panel). Representative images from two individual experiments done in duplicate are shown. Invaded cells were quantified as before and each data point represents mean ± S.D (* indicates p<0.05 compared to LNCaP cells in androgen free medium, n=4) (Right Panel). (c) Plausible mechanism of dual control of p38 MAPK to promote aggressive growth of prostate cancer cells in hypoxia. When subjected to hypoxia, AR can be activated in an androgen independent manner via p38 MAPK and HSP27 and can translocate into the nucleus. On the other hand, activated p38 MAPK can help stabilize HIF-1α and promote HIF-1 mediated gene transcription. Together the active AR and HIF-1 promote aggressive growth and may in due course lead to androgen independent prostate cancer.

Mentions: In an effort to simulate the intermittent hypoxia-reoxygenation conditions that exist in a tumor, LNCaP cells were grown in a cyclic 12h hypoxia and 12h re-oxygenation environment for 3 weeks, passaging the cells as required. Surviving cells (named as hrLNCaP) were collected and equal numbers of cells were seeded in androgen free medium to determine the growth potential of hrLNCaP cells compared to parent LNCaP cells. We observed that continuous cycles of hypoxia re-oxygenation selected for cells with a highly significant faster growth rate compared to the control LNCaP cells (Figure 6a) as determined by MTT assay. hrLNcaP cells also showed higher invasiveness even under normoxic conditions (Figure 6b). The number of cells that invade through the basement matrix was comparable irrespective of the presence or absence of DHT (Figure 6b, Right Panel).


Hypoxia-associated p38 mitogen-activated protein kinase-mediated androgen receptor activation and increased HIF-1alpha levels contribute to emergence of an aggressive phenotype in prostate cancer.

Khandrika L, Lieberman R, Koul S, Kumar B, Maroni P, Chandhoke R, Meacham RB, Koul HK - Oncogene (2009)

Chronic hypoxia- reoxygenation selects for LNCaP cells with aggressive phenotype(a) LNCaP cells were cultured in complete medium and incubated in hypoxia for 12h and re-oxygenated for 12h, for 3 weeks. Equal numbers of surviving cells (hrLNCaP) were seeded in androgen free medium and proliferation was measured by MTT assay over 3 days. Each data point represents mean ± S.D of two individual experiments done in quadruplicate (*** indicates p<0.001, n=8). (b) Equal number of hrLNCaP cells and LNCaP cells were seeded in Boyden chambers in androgen free medium and cells were allowed to invade either in the presence or absence of 10nM DHT. 48h later, invaded cells were stained with Crystal Violet (Left Panel). Representative images from two individual experiments done in duplicate are shown. Invaded cells were quantified as before and each data point represents mean ± S.D (* indicates p<0.05 compared to LNCaP cells in androgen free medium, n=4) (Right Panel). (c) Plausible mechanism of dual control of p38 MAPK to promote aggressive growth of prostate cancer cells in hypoxia. When subjected to hypoxia, AR can be activated in an androgen independent manner via p38 MAPK and HSP27 and can translocate into the nucleus. On the other hand, activated p38 MAPK can help stabilize HIF-1α and promote HIF-1 mediated gene transcription. Together the active AR and HIF-1 promote aggressive growth and may in due course lead to androgen independent prostate cancer.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2651999&req=5

Figure 6: Chronic hypoxia- reoxygenation selects for LNCaP cells with aggressive phenotype(a) LNCaP cells were cultured in complete medium and incubated in hypoxia for 12h and re-oxygenated for 12h, for 3 weeks. Equal numbers of surviving cells (hrLNCaP) were seeded in androgen free medium and proliferation was measured by MTT assay over 3 days. Each data point represents mean ± S.D of two individual experiments done in quadruplicate (*** indicates p<0.001, n=8). (b) Equal number of hrLNCaP cells and LNCaP cells were seeded in Boyden chambers in androgen free medium and cells were allowed to invade either in the presence or absence of 10nM DHT. 48h later, invaded cells were stained with Crystal Violet (Left Panel). Representative images from two individual experiments done in duplicate are shown. Invaded cells were quantified as before and each data point represents mean ± S.D (* indicates p<0.05 compared to LNCaP cells in androgen free medium, n=4) (Right Panel). (c) Plausible mechanism of dual control of p38 MAPK to promote aggressive growth of prostate cancer cells in hypoxia. When subjected to hypoxia, AR can be activated in an androgen independent manner via p38 MAPK and HSP27 and can translocate into the nucleus. On the other hand, activated p38 MAPK can help stabilize HIF-1α and promote HIF-1 mediated gene transcription. Together the active AR and HIF-1 promote aggressive growth and may in due course lead to androgen independent prostate cancer.
Mentions: In an effort to simulate the intermittent hypoxia-reoxygenation conditions that exist in a tumor, LNCaP cells were grown in a cyclic 12h hypoxia and 12h re-oxygenation environment for 3 weeks, passaging the cells as required. Surviving cells (named as hrLNCaP) were collected and equal numbers of cells were seeded in androgen free medium to determine the growth potential of hrLNCaP cells compared to parent LNCaP cells. We observed that continuous cycles of hypoxia re-oxygenation selected for cells with a highly significant faster growth rate compared to the control LNCaP cells (Figure 6a) as determined by MTT assay. hrLNcaP cells also showed higher invasiveness even under normoxic conditions (Figure 6b). The number of cells that invade through the basement matrix was comparable irrespective of the presence or absence of DHT (Figure 6b, Right Panel).

Bottom Line: Our results demonstrate that hypoxia-reoxygenation resulted in increased survival, higher clonogenicity and enhanced invasiveness of these cells.Moreover, hypoxia-reoxygenation was associated with an increased AR activity independent of androgens as well as increased hypoxia inducible factor (HIF-1alpha) levels and activity.We also observed that the activation of p38 mitogen-activated protein (MAP) kinase pathway was an early response to hypoxia, and inhibition of p38 MAP kinase pathway by variety of approaches abolished hypoxia-reoxygenation induced increased AR activity as well as increased survival, clonogenicity and invasiveness.

View Article: PubMed Central - PubMed

Affiliation: Signal Transduction and Molecular Urology Laboratory-Program in Urosciences, Division of Urology, Department of Surgery, School of Medicine, University of Colorado at Denver and Health Sciences Center, Aurora, CO 80045, USA.

ABSTRACT
Androgen receptor (AR) signaling is involved in the development and progression of prostate cancer. Tumor microvasculature contributes to continual exposure of prostate cancer cells to hypoxia-reoxygenation, however, the role of hypoxia-reoxygenation in prostate cancer progression and modulation of AR signaling is not understood. In this study, we evaluated the effects of hypoxia-reoxygenation in LNCaP cells, a line of hormone responsive human prostate cancer cells. Our results demonstrate that hypoxia-reoxygenation resulted in increased survival, higher clonogenicity and enhanced invasiveness of these cells. Moreover, hypoxia-reoxygenation was associated with an increased AR activity independent of androgens as well as increased hypoxia inducible factor (HIF-1alpha) levels and activity. We also observed that the activation of p38 mitogen-activated protein (MAP) kinase pathway was an early response to hypoxia, and inhibition of p38 MAP kinase pathway by variety of approaches abolished hypoxia-reoxygenation induced increased AR activity as well as increased survival, clonogenicity and invasiveness. These results demonstrate a critical role for hypoxia-induced p38 MAP kinase pathway in androgen-independent AR activation in prostate cancer cells, and suggest that hypoxia-reoxygenation may select for aggressive androgen-independent prostate cancer phenotype.

Show MeSH
Related in: MedlinePlus