Limits...
Stromal derived factor-1 (SDF-1/CXCL12) and CXCR4 in renal cell carcinoma metastasis.

Pan J, Mestas J, Burdick MD, Phillips RJ, Thomas GV, Reckamp K, Belperio JA, Strieter RM - Mol. Cancer (2006)

Bottom Line: We demonstrated that CXCR4 was significantly expressed on circulating cytokeratin+ RCC cells from patients with known metastatic RCC.The enhanced CXCR4 expression was mediated through the interaction of the Hypoxia Inducible Factor-1alpha (HIF-1alpha) with the promoter region of the CXCR4 gene.Furthermore, the expression of CXCR4 on human RCC directly correlated with their metastatic ability in vivo in both heterotopic and orthotopic SCID mouse models of human RCC.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Medicine, David Geffen School of Medicine at UCLA, Los Angeles, California, USA. jpan@mednet.ucla.edu

ABSTRACT
Renal cell carcinoma (RCC) is characterized by organ-specific metastases. The chemokine stromal derived factor-1 (SDF-1/CXCL12) and its receptor CXCR4 have been suggested to regulate organ-specific metastasis in various other cancers. On this basis, we hypothesized that the biological axis of CXCL12 via interaction with its receptor, CXCR4, is a major mechanism for RCC metastasis. We demonstrated that CXCR4 was significantly expressed on circulating cytokeratin+ RCC cells from patients with known metastatic RCC. We detected up-regulation of CXCR4 mRNA and protein levels on a human RCC cell line by either knockdown of the von Hippel-Lindau (VHL) tumor suppressor protein, or incubating the cells under hypoxic conditions. The enhanced CXCR4 expression was mediated through the interaction of the Hypoxia Inducible Factor-1alpha (HIF-1alpha) with the promoter region of the CXCR4 gene. Furthermore, the expression of CXCR4 on human RCC directly correlated with their metastatic ability in vivo in both heterotopic and orthotopic SCID mouse models of human RCC. Neutralization of CXCL12 in SCID mice abrogated metastasis of RCC to target organs expressing high levels of CXCL12; without altering tumor cell proliferation, apoptosis, or tumor-associated angiogenesis. Therefore, our data suggest that the CXCL12/CXCR4 biological axis plays an important role in regulating the organ-specific metastasis of RCC.

Show MeSH

Related in: MedlinePlus

Hypoxia and VHL knockdown enhance CXCR4 expression. (A). SN12C-VHL-KD cells were serum-starved and exposed to either normoxia or hypoxia (94% N2, 5% CO2 and 1% O2) for the times indicated. Changes in gene expression were then determined by real-time quantitative PCR. Fold induction represents increases in CXCR4 expression in SN12C-VHL-KD cells compared to SN12C cells transfected with vector control (SN12C-VC). (B). SN12C-P, SN12C-VC and SN12C-VHL-KD cells were treated with either normoxia or hypoxia as described in (A) for the times indicated and then subjected to Western analysis to examine changes in CXCR4 protein levels.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC1636662&req=5

Figure 2: Hypoxia and VHL knockdown enhance CXCR4 expression. (A). SN12C-VHL-KD cells were serum-starved and exposed to either normoxia or hypoxia (94% N2, 5% CO2 and 1% O2) for the times indicated. Changes in gene expression were then determined by real-time quantitative PCR. Fold induction represents increases in CXCR4 expression in SN12C-VHL-KD cells compared to SN12C cells transfected with vector control (SN12C-VC). (B). SN12C-P, SN12C-VC and SN12C-VHL-KD cells were treated with either normoxia or hypoxia as described in (A) for the times indicated and then subjected to Western analysis to examine changes in CXCR4 protein levels.

Mentions: Recently, the expression of CXCR4 on RCC and NSCLC tumor cells was shown to be regulated by hypoxia and the VHL/HIF-1α pathway [33-35]. To further characterize the role of hypoxia and/or VHL/HIF-1α in the regulation of the expression of CXCR4 on RCC cells and to determine whether this chemokine receptor is a critical factor for promoting RCC metastases, we next exposed SN12C-P, SN12C-VC and SN12C-VHL-KD cells to normoxia or hypoxia for 2, 4, 8, and 24 hours and isolated mRNA and protein for quantitative TaqMan RT-PCR and Western blot analysis of CXCR4, respectively. As shown in Fig 2A, SN12C-VHL-KD cells demonstrated a marked increase in CXCR4 gene expression under both normoxic and hypoxic conditions, as compared to SN12C-VC. Similar results were found when comparing the SN12C-VHL-KD to SN12C-P (data not shown). Total cellular levels of CXCR4 protein paralleled the expression of mRNA in SN12C-VHL-KD cells with significantly higher levels of CXCR4 protein found under both conditions of normoxia and hypoxia, as compared to either SN12C-P or SN12C-VC cell lines (Fig. 2B). These latter two cell lines demonstrated markedly increased CXCR4 protein levels only under hypoxic conditions (Fig. 2B). CXCR4 cell surface expression was seen by FACS analysis and correlated with the results obtained from Western blot analysis (data not shown).


Stromal derived factor-1 (SDF-1/CXCL12) and CXCR4 in renal cell carcinoma metastasis.

Pan J, Mestas J, Burdick MD, Phillips RJ, Thomas GV, Reckamp K, Belperio JA, Strieter RM - Mol. Cancer (2006)

Hypoxia and VHL knockdown enhance CXCR4 expression. (A). SN12C-VHL-KD cells were serum-starved and exposed to either normoxia or hypoxia (94% N2, 5% CO2 and 1% O2) for the times indicated. Changes in gene expression were then determined by real-time quantitative PCR. Fold induction represents increases in CXCR4 expression in SN12C-VHL-KD cells compared to SN12C cells transfected with vector control (SN12C-VC). (B). SN12C-P, SN12C-VC and SN12C-VHL-KD cells were treated with either normoxia or hypoxia as described in (A) for the times indicated and then subjected to Western analysis to examine changes in CXCR4 protein levels.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1636662&req=5

Figure 2: Hypoxia and VHL knockdown enhance CXCR4 expression. (A). SN12C-VHL-KD cells were serum-starved and exposed to either normoxia or hypoxia (94% N2, 5% CO2 and 1% O2) for the times indicated. Changes in gene expression were then determined by real-time quantitative PCR. Fold induction represents increases in CXCR4 expression in SN12C-VHL-KD cells compared to SN12C cells transfected with vector control (SN12C-VC). (B). SN12C-P, SN12C-VC and SN12C-VHL-KD cells were treated with either normoxia or hypoxia as described in (A) for the times indicated and then subjected to Western analysis to examine changes in CXCR4 protein levels.
Mentions: Recently, the expression of CXCR4 on RCC and NSCLC tumor cells was shown to be regulated by hypoxia and the VHL/HIF-1α pathway [33-35]. To further characterize the role of hypoxia and/or VHL/HIF-1α in the regulation of the expression of CXCR4 on RCC cells and to determine whether this chemokine receptor is a critical factor for promoting RCC metastases, we next exposed SN12C-P, SN12C-VC and SN12C-VHL-KD cells to normoxia or hypoxia for 2, 4, 8, and 24 hours and isolated mRNA and protein for quantitative TaqMan RT-PCR and Western blot analysis of CXCR4, respectively. As shown in Fig 2A, SN12C-VHL-KD cells demonstrated a marked increase in CXCR4 gene expression under both normoxic and hypoxic conditions, as compared to SN12C-VC. Similar results were found when comparing the SN12C-VHL-KD to SN12C-P (data not shown). Total cellular levels of CXCR4 protein paralleled the expression of mRNA in SN12C-VHL-KD cells with significantly higher levels of CXCR4 protein found under both conditions of normoxia and hypoxia, as compared to either SN12C-P or SN12C-VC cell lines (Fig. 2B). These latter two cell lines demonstrated markedly increased CXCR4 protein levels only under hypoxic conditions (Fig. 2B). CXCR4 cell surface expression was seen by FACS analysis and correlated with the results obtained from Western blot analysis (data not shown).

Bottom Line: We demonstrated that CXCR4 was significantly expressed on circulating cytokeratin+ RCC cells from patients with known metastatic RCC.The enhanced CXCR4 expression was mediated through the interaction of the Hypoxia Inducible Factor-1alpha (HIF-1alpha) with the promoter region of the CXCR4 gene.Furthermore, the expression of CXCR4 on human RCC directly correlated with their metastatic ability in vivo in both heterotopic and orthotopic SCID mouse models of human RCC.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Medicine, David Geffen School of Medicine at UCLA, Los Angeles, California, USA. jpan@mednet.ucla.edu

ABSTRACT
Renal cell carcinoma (RCC) is characterized by organ-specific metastases. The chemokine stromal derived factor-1 (SDF-1/CXCL12) and its receptor CXCR4 have been suggested to regulate organ-specific metastasis in various other cancers. On this basis, we hypothesized that the biological axis of CXCL12 via interaction with its receptor, CXCR4, is a major mechanism for RCC metastasis. We demonstrated that CXCR4 was significantly expressed on circulating cytokeratin+ RCC cells from patients with known metastatic RCC. We detected up-regulation of CXCR4 mRNA and protein levels on a human RCC cell line by either knockdown of the von Hippel-Lindau (VHL) tumor suppressor protein, or incubating the cells under hypoxic conditions. The enhanced CXCR4 expression was mediated through the interaction of the Hypoxia Inducible Factor-1alpha (HIF-1alpha) with the promoter region of the CXCR4 gene. Furthermore, the expression of CXCR4 on human RCC directly correlated with their metastatic ability in vivo in both heterotopic and orthotopic SCID mouse models of human RCC. Neutralization of CXCL12 in SCID mice abrogated metastasis of RCC to target organs expressing high levels of CXCL12; without altering tumor cell proliferation, apoptosis, or tumor-associated angiogenesis. Therefore, our data suggest that the CXCL12/CXCR4 biological axis plays an important role in regulating the organ-specific metastasis of RCC.

Show MeSH
Related in: MedlinePlus