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Primary research
Jun Zhang
Department of Morphological Experiment Center, Medical College of Yanbian University, Yanji City (133000), Jilin Province, China.
Corresponding Author
ORCiD: https://orcid.org/0000-0001-7111-0205
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: Forkhead box protein H1 (FOXH1) is upregulated in a variety of cancer types but its expression patterns and specific functions in lung cancer are unclear at present. The main objective of the present study was to establish whether FOXH1 plays a role in regulation of lung cancer progression.
Methods: The TCGA and Kaplan-Meier plotter dataset was analyzed for possible association between FOXH1 expressions in lung cancer tissues and patient prognosis. A549 and PC9 cells were transfected with shRNA targeting FOXH1 mRNA. The Cell Counting Kit-8 (CCK8), plate clone formation, soft agar, wound healing, transwell invasion and flow cytometry assay were performed to detect the cell proliferation, migration and invasion in lung cancer cells. Tumorigenicity was examined in mouse model system. Western blot analysis was used to detect FOXH1, Matrix metalloproteinase 2 (MMP2), Vimentin, N-cadherin, E-cadherin, Snail, Slug, p-GSK-3β, β-catenin and Cyclin D1. The β-catenin activity was measured by luciferase reporter system assay.
Results: FOXH1 expression in the lung cancer was determined using the TCGA and Kaplan-Meier plotter databases. Higher expression of FOXH1 was observed in tumor tissue relative to normal tissue, which was associated with reduced overall survival. FOXH1 knockdown resulted in significant inhibition of the proliferation, the cell cycle, the migration as well as the invasion of lung cancer cells. Then, we confirmed that FOXH1 knockdown can significantly slow down tumor growth and tumor cell proliferation in vivo using the mouse model. It also significantly reduced the migration and invasion capabilities of lung cancer cells. Using Western blot analyses, we found that FOXH1 depletion inhibited the epithelial-mesenchymal transition of lung cancer cells through downregulation of the mesenchymal cell markers Snail, Slug, MMP2, N-cadherin, and Vimentin, and upregulation of the epithelial marker E-cadherin. Moreover, FOXH1 knockdown significantly downregulated β-catenin and its downstream targets, such as p-GSK-3β and cyclin D1, and also led to direct suppression of β-catenin activity, as determined by the luciferase reporter system assay.
Conclusion: In conclusion, FOXH1 promotes proliferation, migration, and invasion of lung cancer cells via regulation of Wnt/β-catenin signaling. FOXH1 is a prognostic marker and a potential therapeutic target for lung cancer treatment.
Keywords
FOXH1, Lung cancer, proliferation, EMT, β-catenin
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On 25 Feb, 2021
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On 09 Feb, 2021
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Editorial decision: Revise Before Review
On 25 Jan, 2021
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Subject Areas
Cancer Biology
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This preprint is under consideration at Cancer Cell International. A preprint is a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Learn more about In Review
Primary research
Jun Zhang
Department of Morphological Experiment Center, Medical College of Yanbian University, Yanji City (133000), Jilin Province, China.
Corresponding Author
ORCiD: https://orcid.org/0000-0001-7111-0205
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Under Revision
Version (no preprint)
Posted 28 Feb, 2021
Editorial decision: Revise Before Review
On 28 Feb, 2021
Editor assigned
On 25 Feb, 2021
Submission checks complete
On 25 Feb, 2021
Editor invited
On 25 Feb, 2021
This version was not preprinted
Version (no preprint)
Posted 10 Feb, 2021
Editorial decision: Revise Before Review
On 10 Feb, 2021
Editor assigned
On 09 Feb, 2021
Submission checks complete
On 09 Feb, 2021
Editor invited
On 09 Feb, 2021
This version was not preprinted
Version 1
Posted 01 Feb, 2021
No community comments so far
Editorial decision: Revise Before Review
On 25 Jan, 2021
Editor assigned
On 24 Jan, 2021
Submission checks complete
On 24 Jan, 2021
Editor invited
On 24 Jan, 2021
First submitted
On 22 Jan, 2021
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Cancer Biology
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: Forkhead box protein H1 (FOXH1) is upregulated in a variety of cancer types but its expression patterns and specific functions in lung cancer are unclear at present. The main objective of the present study was to establish whether FOXH1 plays a role in regulation of lung cancer progression.
Methods: The TCGA and Kaplan-Meier plotter dataset was analyzed for possible association between FOXH1 expressions in lung cancer tissues and patient prognosis. A549 and PC9 cells were transfected with shRNA targeting FOXH1 mRNA. The Cell Counting Kit-8 (CCK8), plate clone formation, soft agar, wound healing, transwell invasion and flow cytometry assay were performed to detect the cell proliferation, migration and invasion in lung cancer cells. Tumorigenicity was examined in mouse model system. Western blot analysis was used to detect FOXH1, Matrix metalloproteinase 2 (MMP2), Vimentin, N-cadherin, E-cadherin, Snail, Slug, p-GSK-3β, β-catenin and Cyclin D1. The β-catenin activity was measured by luciferase reporter system assay.
Results: FOXH1 expression in the lung cancer was determined using the TCGA and Kaplan-Meier plotter databases. Higher expression of FOXH1 was observed in tumor tissue relative to normal tissue, which was associated with reduced overall survival. FOXH1 knockdown resulted in significant inhibition of the proliferation, the cell cycle, the migration as well as the invasion of lung cancer cells. Then, we confirmed that FOXH1 knockdown can significantly slow down tumor growth and tumor cell proliferation in vivo using the mouse model. It also significantly reduced the migration and invasion capabilities of lung cancer cells. Using Western blot analyses, we found that FOXH1 depletion inhibited the epithelial-mesenchymal transition of lung cancer cells through downregulation of the mesenchymal cell markers Snail, Slug, MMP2, N-cadherin, and Vimentin, and upregulation of the epithelial marker E-cadherin. Moreover, FOXH1 knockdown significantly downregulated β-catenin and its downstream targets, such as p-GSK-3β and cyclin D1, and also led to direct suppression of β-catenin activity, as determined by the luciferase reporter system assay.
Conclusion: In conclusion, FOXH1 promotes proliferation, migration, and invasion of lung cancer cells via regulation of Wnt/β-catenin signaling. FOXH1 is a prognostic marker and a potential therapeutic target for lung cancer treatment.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
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