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N. Kang
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P3.01 - Poster Session with Presenters Present (ID 469)
- Event: WCLC 2016
- Type: Poster Presenters Present
- Track: Biology/Pathology
- Presentations: 3
- Moderators:
- Coordinates: 12/07/2016, 14:30 - 15:45, Hall B (Poster Area)
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P3.01-056 - Association of Angiogenesis, EMT and Stem Cell Characteristics Using Hypoxic Stress in Lung Cancer (ID 4220)
14:30 - 14:30 | Author(s): N. Kang
- Abstract
Background:
Hypoxia, a major phenomenon in solid tumors, can promote the metastatic potential of tumor cells which is associated with chemoresistance and poor prognosis. It was reported that various angiogenesis factors including VEGF and HIF, were associated in cancer development and progression by hypoxia. In addition, both epithelial-mesenchymal transition (EMT) and cancer stem cells play an important role in malignant progression in many human tumors. We investigated the effect of hypoxic stress on the angiogenesis, EMT and stemness acquisition in lung cancer.
Methods:
Normal lung cell (BEAS-2B) and lung cancer cell lines (A549, H292, H226 and H460) were incubated in either normoxic or hypoxic (below 1% O~2~) conditions. For transcriptome analysis, mRNA of BEAS-2B and A549 cell lines were analyzed using next-generation sequencing (HiSeq 2500 system). For further validation, angiogenesis markers were analyzed by western blotting. EMT was assessed with western blotting, wound healing assay and Matrigel invasion assay, and stem cell characteristics were assessed with RT-PCR, immunostaining, soft agar colony formation assay, sphere formation assay and in vivo mice tumor model.
Results:
Next-generation sequencing revealed significant changes in the expression of angiogenesis, EMT and stem cell markers after hypoxic stress. Among the angiogenesis markers, VEGF and HIF-2α were increased. EMT markers related in hypoxia showed decrease in E-cadherin and increase in fibronectin, vimentin, N-cadherin, α-SMA, Snail, Slug, ZEB1 and ZEB2. Stem cell markers such as CXCR4, Oct4 and Nanog were increased at least one lung cancer cell line in hypoxic condition compared with in normoxic condition. Functional assays for EMT and stemness acquisition indicated that hypoxic stress increased wound healing, Matrigel invasion, sphere formation and in vivo mice tumor formation.
Conclusion:
These results suggest that hypoxia induces angiogenesis markers expression which is associated with EMT and stemness acquisition in lung cancer.
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P3.01-057 - TGF-β Induced EMT and Stemness Characteristics in Lung Cancer (ID 4221)
14:30 - 14:30 | Author(s): N. Kang
- Abstract
Background:
Transforming growth factor-β (TGF-β) is known to inhibit cell growth in benign cells but promotes tumor invasion and metastasis by inducing an epithelial-mesenchymal transition (EMT). EMT is a differentiation switch through which epithelial cells differentiate into mesenchymal cells. It occurs in the process of tissue morphogenesis during development, wound repair and cancer progression in adult tissues. EMT is often associated with acquisition of stem-like characteristics. In this study, we investigated whether EMT induced by TGF-β could acquire stem-like characteristics in lung cancer.
Methods:
Human normal epithelial (BEAS-2B) and cancer (A549, H292, H226 and H460) cell lines were incubated with 10 ng/ml of TGF-β for 3 days. Transcriptome and methylation analysis of BEAS-2B and A549 cells treated with TGF-β were performed by using next-generation sequencing (HiSeq 2500 system). Western blotting was performed to analyze the expression of epithelial marker (E-cadherin) and mesenchymal markers (fibronectin, vimentin, N-cadherin and α-SMA). RT-PCR was performed to analyze the expression of variable stem cell markers (CD44, CD133, CXCR4, ABCG2, CD117, ALDH1A1, EpCAM, CD90, Oct4, Nanog, SOX2, SSEA4, and CD166). Wound healing assay, Matrigel invasion assay and sphere formation assay were used to assess functional characteristics of EMT and stemness acquisition.
Results:
Next-generation sequencing revealed significant changes in the expression of stem cell markers, CD44, ALDH1A1 and CD90 in both BEAS-2B and A549 cells. The changes in the expression of EMT and stem cell markers induced by TGF-β were variable according to lung cell lines. Except for H460 cell line, lung cell lines showed at least one or more increased stem cell markers expression with TGF-β. Functional analysis revealed increased wound healing, Matrigel invasion and sphere formation after TGF-β treatment
Conclusion:
TGF-β induced EMT was associated with acquisition of stem-like characteristics. Various expression patterns of stem cell marker were observed according to different lung cancer cell lines.
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P3.01-058 - Demethylation of CXCR4 and Stemness Acquisition in Lung Cancer (ID 4222)
14:30 - 14:30 | Author(s): N. Kang
- Abstract
Background:
As a cancer stem cell marker, CXCR4 has been known to be closely associated with cell survival and stemness acquisition. Previous studies reported that the level of CXCR4 is increased after hypoxic condition in several types of cancer. However, the mechanism of the increased CXCR4 expression has not been well understood. We investigated whether aberrant promoter demethylation could induce CXCR4 activation by using hypoxic stress in lung cancer.
Methods:
Human normal lung cell (BEAS-2B) and lung cancer cell lines (A549, H292, H226 and H460) were incubated under hypoxic condition. Transcriptome and methylation analysis using next-generation sequencing were performed by HiSeq 2500 system. For further validation, CXCR4 expression was analyzed by RT-PCR and western blotting. To determine whether CXCR4 is reactivated, cell lines were treated with a DNA methyltransferase inhibitor (AZA). Hypoxia-induced DNA demethylation was identified by methylation-specific PCR and bisulfite sequencing. Stem cell characteristics were assessed by sphere formation assay and in vivo mice tumor model.
Results:
Next-generation sequencing results revealed that CXCR4 expression was increased after hypoxic condition, whereas CXCR4 methylation was reduced. CXCR4 was activated by either hypoxic condition and treatment with AZA. MSP showed decreased CXCR4 promoter methylation in hypoxic condition compared with normoxic condition. Functional stem cell assay indicated that hypoxic stress increased sphere formation and in vivo mice tumor formation.
Conclusion:
These results suggest that hypoxia induces stem cell characteristics which are related with CXCR4 reactivation by promoter demethylation.
