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Nuplazid suppresses esophageal squamous cell carcinoma growth in vitro and in vivo by targeting PAK4. Br J Cancer 2021; 126:1037-1046. [PMID: 34912075 PMCID: PMC8980085 DOI: 10.1038/s41416-021-01651-z] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2021] [Revised: 10/31/2021] [Accepted: 11/22/2021] [Indexed: 11/09/2022] Open
Abstract
Background Due to the high recurrence and low 5-year survival rates of esophageal squamous cell carcinoma (ESCC) after treatment, the discovery of novel drugs for recurrence chemoprevention is of particular importance. Methods We screened the FDA-approved drug library and found that Nuplazid, an atypical antipsychotic that acts as an effective 5-HT 2 A receptor inverse agonist, could potentially exert anticancer effects in vitro and in vivo on ESCC. Results Pull-down results indicated that Nuplazid binds with p21-activated kinase 4 (PAK4), and a kinase assay showed that Nuplazid strongly suppressed PAK4 kinase activity. Moreover, Nuplazid exhibited inhibitory effects on ESCC in vivo. Conclusions Our findings indicate that Nuplazid can suppress ESCC progression through targeting PAK4.
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Huang H, Xue Q, Du X, Cui J, Wang J, Cheng D, Li J, Zheng Y, Huang G, Zhang K, Liu K, Lu J, Zhao J, Chen X, Dong Z, Li X. p21-activated kinase 4 promotes the progression of esophageal squamous cell carcinoma by targeting LASP1. Mol Carcinog 2020; 60:38-50. [PMID: 33289209 PMCID: PMC7756368 DOI: 10.1002/mc.23269] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/26/2019] [Revised: 08/25/2020] [Accepted: 11/19/2020] [Indexed: 12/30/2022]
Abstract
Esophageal squamous cell carcinoma (ESCC) is one of the most common malignant tumors of the digestive tract in humans. Several studies have indicated that PAK4 is associated with the risk of ESCC and may be a potential druggable kinase for ESCC treatment. However, the underlying mechanism remains largely unknown. The aim of our study is to identify the functional role of PAK4 in ESCC. To determine the expression of PAK4 in ESCC, Western blot analysis and immunohistochemistry were performed, and the results showed that PAK4 is significantly upregulated in ESCC tissues and cell lines compared with normal controls and normal esophageal epithelial cell line. To further investigate the role of PAK4 in ESCC, cell viability assays, anchorage-independent cell growth assays, wound healing assays, cellular invasion assays, in vivo xenograft mouse models, and metastasis assays were conducted, and the results showed that PAK4 can significantly facilitate ESCC proliferation and metastasis in vitro and in vivo. To determine the potential target of PAK4 in ESCC progression, a pull-down assay was performed, and the results showed that LASP1 may be a potential target of PAK4. An immunoprecipitation assay and confocal microscopy analysis confirmed that PAK4 can bind to and colocalize with LASP1 in vitro and in cells. Notably, rescue experiments further illustrated the mechanistic network of PAK4/LASP1. Our research reveals the oncogenic roles of PAK4 in ESCC and preliminarily elucidates the mechanistic network of PAK4/LASP1 in ESCC.
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Affiliation(s)
- Hui Huang
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
| | - Qianqian Xue
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- Department of Public HealthNanshi Hospital of NanyangNanyangHenanChina
| | - Xiaoge Du
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- China‐US (Henan) Hormel Cancer InstituteZhengzhouHenanChina
- Department of NursingHenan Health School of Medicine and PharmacyPingdingshanHenanChina
| | - Jie Cui
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
| | - Jing Wang
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- China‐US (Henan) Hormel Cancer InstituteZhengzhouHenanChina
| | - Dan Cheng
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- China‐US (Henan) Hormel Cancer InstituteZhengzhouHenanChina
| | - Jiaqiong Li
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- China‐US (Henan) Hormel Cancer InstituteZhengzhouHenanChina
| | - Yaqiu Zheng
- China‐US (Henan) Hormel Cancer InstituteZhengzhouHenanChina
| | - Guojing Huang
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
| | - Keke Zhang
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- China‐US (Henan) Hormel Cancer InstituteZhengzhouHenanChina
| | - Kangdong Liu
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- China‐US (Henan) Hormel Cancer InstituteZhengzhouHenanChina
- Collaborative Innovation Center of Henan Province for Cancer ChemopreventionZhengzhouHenanChina
- State Key Laboratory of Esophageal Cancer Prevention and Treatment, Zhengzhou UniversityZhengzhouHenanChina
| | - Jing Lu
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- Collaborative Innovation Center of Henan Province for Cancer ChemopreventionZhengzhouHenanChina
- State Key Laboratory of Esophageal Cancer Prevention and Treatment, Zhengzhou UniversityZhengzhouHenanChina
| | - Jimin Zhao
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- Collaborative Innovation Center of Henan Province for Cancer ChemopreventionZhengzhouHenanChina
- State Key Laboratory of Esophageal Cancer Prevention and Treatment, Zhengzhou UniversityZhengzhouHenanChina
| | - Xinhuan Chen
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- Collaborative Innovation Center of Henan Province for Cancer ChemopreventionZhengzhouHenanChina
- State Key Laboratory of Esophageal Cancer Prevention and Treatment, Zhengzhou UniversityZhengzhouHenanChina
| | - Ziming Dong
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- Collaborative Innovation Center of Henan Province for Cancer ChemopreventionZhengzhouHenanChina
- State Key Laboratory of Esophageal Cancer Prevention and Treatment, Zhengzhou UniversityZhengzhouHenanChina
| | - Xiang Li
- Department of Pathophysiology, School of Basic Medical SciencesZhengzhou UniversityZhengzhouHenanChina
- China‐US (Henan) Hormel Cancer InstituteZhengzhouHenanChina
- Collaborative Innovation Center of Henan Province for Cancer ChemopreventionZhengzhouHenanChina
- State Key Laboratory of Esophageal Cancer Prevention and Treatment, Zhengzhou UniversityZhengzhouHenanChina
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Xie Y, Zhang J, Lu B, Bao Z, Zhao J, Lu X, Wei Y, Yao K, Jiang Y, Yuan Q, Zhang X, Li B, Chen X, Dong Z, Liu K. Mefloquine Inhibits Esophageal Squamous Cell Carcinoma Tumor Growth by Inducing Mitochondrial Autophagy. Front Oncol 2020; 10:1217. [PMID: 32850358 PMCID: PMC7400730 DOI: 10.3389/fonc.2020.01217] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2020] [Accepted: 06/15/2020] [Indexed: 12/12/2022] Open
Abstract
Esophageal squamous cell carcinoma (ESCC) has a worldwide impact on human health, due to its high incidence and mortality. Therefore, identifying compounds to increase patients' survival rate is urgently needed. Mefloquine (MQ) is an FDA-approved anti-malarial drug, which has been reported to inhibit cellular proliferation in several cancers. However, the anti-tumor activities of the drug have not yet been completely defined. In this study, mass spectrometry was employed to profile proteome changes in ESCC cells after MQ treatment. Sub-cellular localization and gene ontology term enrichment analysis suggested that MQ treatment mainly affect mitochondria. The KEGG pathway enrichment map of down-regulated pathways and Venn diagram indicated that all of the top five down regulated signaling pathways contain four key mitochondrial proteins (succinate dehydrogenase complex subunit C (SDHC), succinate dehydrogenase complex subunit D, mitochondrially encoded cytochrome c oxidase III and NADH: ubiquinone oxidoreductase subunit V3). Meanwhile, mitochondrial autophagy was observed in MQ-treated KYSE150 cells. More importantly, patient-derived xenograft mouse models of ESCC with SDHC high expression were more sensitive to MQ treatment than low SDHC-expressing xenografts. Taken together, mefloquine inhibits ESCC tumor growth by inducing mitochondrial autophagy and SDHC plays a vital role in MQ-induced anti-tumor effect on ESCC.
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Affiliation(s)
- Yifei Xie
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China
| | - Jing Zhang
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China
| | - Bingbing Lu
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China
| | - Zhuo Bao
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China
| | - Jimin Zhao
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China.,Henan Provincial Cooperative Innovation Center for Cancer Chemoprevention, Zhengzhou, China
| | - Xianyu Lu
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China
| | - Yaxing Wei
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China
| | - Ke Yao
- China-US (Henan) Hormel Cancer Institute, Zhengzhou, China
| | - Yanan Jiang
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China
| | - Qiang Yuan
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China
| | - Xiaofan Zhang
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China
| | - Bo Li
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China
| | - Xinhuan Chen
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China.,Henan Provincial Cooperative Innovation Center for Cancer Chemoprevention, Zhengzhou, China
| | - Zigang Dong
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China.,China-US (Henan) Hormel Cancer Institute, Zhengzhou, China.,State Key Laboratory of Esophageal Cancer Prevention and Treatment, Zhengzhou, China.,Cancer Chemoprevention International Collaboration Laboratory, Zhengzhou, China
| | - Kangdong Liu
- Department of Pathophysiology, School of Basic Medical Sciences, AMS, Zhengzhou University, Zhengzhou, China.,Henan Provincial Cooperative Innovation Center for Cancer Chemoprevention, Zhengzhou, China.,China-US (Henan) Hormel Cancer Institute, Zhengzhou, China.,State Key Laboratory of Esophageal Cancer Prevention and Treatment, Zhengzhou, China.,Cancer Chemoprevention International Collaboration Laboratory, Zhengzhou, China
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4
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Cheng YW, Chen YM, Zhao QQ, Zhao X, Wu YR, Chen DZ, Liao LD, Chen Y, Yang Q, Xu LY, Li EM, Xu JZ. Long Read Single-Molecule Real-Time Sequencing Elucidates Transcriptome-Wide Heterogeneity and Complexity in Esophageal Squamous Cells. Front Genet 2019; 10:915. [PMID: 31636653 PMCID: PMC6787290 DOI: 10.3389/fgene.2019.00915] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2019] [Accepted: 08/29/2019] [Indexed: 02/05/2023] Open
Abstract
Esophageal squamous cell carcinoma is a leading cause of cancer death. Mapping the transcriptional landscapes such as isoforms, fusion transcripts, as well as long noncoding RNAs have played a central role to understand the regulating mechanism during malignant processes. However, canonical methods such as short-read RNA-seq are difficult to define the entire polyadenylated RNA molecules. Here, we combined single-molecule real-time sequencing with RNA-seq to generate high-quality long reads and to survey the transcriptional program in esophageal squamous cells. Compared with the recent annotations of human transcriptome (Ensembl 38 release 91), single-molecule real-time data identified many unannotated transcripts, novel isoforms of known genes and an expanding repository of long intergenic noncoding RNAs (lincRNAs). By integrating with annotation of lincRNA catalog, 1,521 esophageal-cancer-specific lincRNAs were defined from single-molecule real-time reads. Kyoto Encyclopedia of Genes and Genomes enrichment analysis indicated that these lincRNAs and their target genes are involved in a variety of cancer signaling pathways. Isoform usage analysis revealed the shifted alternative splicing patterns, which can be recaptured from clinical samples or supported by previous studies. Utilizing vigorous searching criteria, we also detected multiple transcript fusions, which are not documented in current gene fusion database or readily identified from RNA-seq reads. Two novel fusion transcripts were verified based on real-time PCR and Sanger sequencing. Overall, our long-read single-molecule sequencing largely expands current understanding of full-length transcriptome in esophageal cells and provides novel insights on the transcriptional diversity during oncogenic transformation.
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Affiliation(s)
- Yin-Wei Cheng
- The Key Laboratory of Molecular Biology for High Cancer Incidence Coastal Chaoshan Area, Shantou University Medical College, Shantou, China
- Department of Biochemistry and Molecular Biology, Shantou University Medical College, Shantou, China
- Computational Systems Biology Lab, Department of Bioinformatics, Shantou University Medical College (SUMC), Shantou, China
| | - Yun-Mei Chen
- Tianjin Novogene Bioinformatics Technology Co., Ltd, Tianjin, China
| | - Qian-Qian Zhao
- Computational Systems Biology Lab, Department of Bioinformatics, Shantou University Medical College (SUMC), Shantou, China
| | - Xing Zhao
- Computational Systems Biology Lab, Department of Bioinformatics, Shantou University Medical College (SUMC), Shantou, China
| | - Ya-Ru Wu
- Computational Systems Biology Lab, Department of Bioinformatics, Shantou University Medical College (SUMC), Shantou, China
| | - Dan-Ze Chen
- Computational Systems Biology Lab, Department of Bioinformatics, Shantou University Medical College (SUMC), Shantou, China
| | - Lian-Di Liao
- The Key Laboratory of Molecular Biology for High Cancer Incidence Coastal Chaoshan Area, Shantou University Medical College, Shantou, China
- China Institute of Oncologic Pathology, Shantou University Medical College, Shantou, China
| | - Yang Chen
- The Key Laboratory of Molecular Biology for High Cancer Incidence Coastal Chaoshan Area, Shantou University Medical College, Shantou, China
- Department of Biochemistry and Molecular Biology, Shantou University Medical College, Shantou, China
| | - Qian Yang
- The Key Laboratory of Molecular Biology for High Cancer Incidence Coastal Chaoshan Area, Shantou University Medical College, Shantou, China
- Department of Biochemistry and Molecular Biology, Shantou University Medical College, Shantou, China
| | - Li-Yan Xu
- The Key Laboratory of Molecular Biology for High Cancer Incidence Coastal Chaoshan Area, Shantou University Medical College, Shantou, China
- China Institute of Oncologic Pathology, Shantou University Medical College, Shantou, China
- *Correspondence: Li-Yan Xu, ; En-Min Li, ; Jian-Zhen Xu,
| | - En-Min Li
- The Key Laboratory of Molecular Biology for High Cancer Incidence Coastal Chaoshan Area, Shantou University Medical College, Shantou, China
- Department of Biochemistry and Molecular Biology, Shantou University Medical College, Shantou, China
- *Correspondence: Li-Yan Xu, ; En-Min Li, ; Jian-Zhen Xu,
| | - Jian-Zhen Xu
- The Key Laboratory of Molecular Biology for High Cancer Incidence Coastal Chaoshan Area, Shantou University Medical College, Shantou, China
- Computational Systems Biology Lab, Department of Bioinformatics, Shantou University Medical College (SUMC), Shantou, China
- *Correspondence: Li-Yan Xu, ; En-Min Li, ; Jian-Zhen Xu,
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Sun J, Yan J, Yuan X, Yang R, Dan T, Wang X, Kong G, Gao S. A computationally constructed ceRNA interaction network based on a comparison of the SHEE and SHEEC cell lines. Cell Mol Biol Lett 2016; 21:21. [PMID: 28536623 PMCID: PMC5415789 DOI: 10.1186/s11658-016-0022-0] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/17/2016] [Accepted: 09/10/2016] [Indexed: 12/16/2022] Open
Abstract
Long non-coding RNAs (lncRNAs) play critical and complicated roles in the regulation of various biological processes, including chromatin modification, transcription and post-transcriptional processing. Interestingly, some lncRNAs serve as miRNA "sponges" that inhibit interaction with miRNA targets in post-transcriptional regulation. We constructed a putative competing endogenous RNA (ceRNA) network by integrating lncRNA, miRNA and mRNA expression based on high-throughput RNA sequencing and microarray data to enable a comparison of the SHEE and SHEEC cell lines. Using Targetscan and miRanda bioinformatics algorithms and miRTarbase microRNA-target interactions database, we established that 51 miRNAs sharing 13,623 MREs with 2260 genes and 82 lncRNAs were involved in this ceRNA network. Through a biological function analysis, the ceRNA network appeared to be primarily involved in cell proliferation, apoptosis, the cell cycle, invasion and metastasis. Functional pathway analyses demonstrated that the ceRNA network potentially modulated multiple signaling pathways, such as the MAPK, Ras, HIF-1, Rap1, and PI3K/Akt signaling pathways. These results might provide new clues to better understand the regulation of the ceRNA network in cancer.
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Affiliation(s)
- Jiachun Sun
- Department of Oncology, Cancer Institute, First Affiliated Hospital of Henan University of Science and Technology, Luoyang, People’s Republic of China
| | - Junqiang Yan
- Department of Neurology, Cancer Institute, First Affiliated Hospital of Henan University of Science and Technology, Luoyang, People’s Republic of China
| | - Xiaozhi Yuan
- Department of Oncology, Cancer Institute, First Affiliated Hospital of Henan University of Science and Technology, Luoyang, People’s Republic of China
| | - Ruina Yang
- Department of Oncology, Cancer Institute, First Affiliated Hospital of Henan University of Science and Technology, Luoyang, People’s Republic of China
| | - Tanyou Dan
- Department of Oncology, Cancer Institute, First Affiliated Hospital of Henan University of Science and Technology, Luoyang, People’s Republic of China
| | - Xinshuai Wang
- Department of Oncology, Cancer Institute, First Affiliated Hospital of Henan University of Science and Technology, Luoyang, People’s Republic of China
| | - Guoqiang Kong
- Department of Oncology, Cancer Institute, First Affiliated Hospital of Henan University of Science and Technology, Luoyang, People’s Republic of China
| | - Shegan Gao
- Department of Oncology, Cancer Institute, First Affiliated Hospital of Henan University of Science and Technology, Luoyang, People’s Republic of China
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Abstract
Transforming growth factor-beta (TGF-beta) represents a large family of growth and differentiation factors that mobilize complex signaling networks to regulate cellular differentiation, proliferation, motility, adhesion, and apoptosis. TGF-beta signaling is tightly regulated by multiple complex mechanisms, and its deregulation plays a key role in the progression of many forms of cancer. Upon ligand binding, TGF-beta signals are transduced by Smad proteins, which in turn are tightly dependent on modulation by adaptor proteins such as embryonic liver fodrin, Smad anchor for receptor activation, filamin, and crkl. A further layer of regulation is imposed by ubiquitin-mediated targeting and proteasomal degradation of specific components of the TGF-beta signaling pathway. This review focuses on the ubiquitinators that regulate TGF-beta signaling and the association of these ubiquitin ligases with various forms of cancer. Delineating the role of ubiquitinators in the TGF-beta signaling pathway could yield powerful novel therapeutic targets for designing new cancer treatments.
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Affiliation(s)
- Eric Glasgow
- Laboratory of Cancer Genetics, Digestive Diseases, and GI Developmental Biology, Department of Surgery, Medicine and Lombardi Cancer Center, Georgetown University Medical Center, Washington, DC 20007, USA.
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N/A, 齐 义, 王 立. N/A. Shijie Huaren Xiaohua Zazhi 2005; 13:1317-1321. [DOI: 10.11569/wcjd.v13.i11.1317] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/26/2023] Open
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Xiong XD, Li EM, Xu LY, Chen HB, Chen L, Cai WJ, Han YL, Shen ZY, Zeng Y. Separation and identification of differentially expressed nuclear matrix proteins between human esophageal immortalized and carcinomatous cell lines. World J Gastroenterol 2003; 9:2143-8. [PMID: 14562366 PMCID: PMC4656451 DOI: 10.3748/wjg.v9.i10.2143] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To separate and identify differentially expressed nuclear matrix proteins (NMPs) between the immortalized human esophageal epithelial cell line (SHEE) and the malignantly transformed esophageal carcinoma cell line (SHEEC), and to provide new ways for finding specific markers and the pathogenesis of esophageal carcinoma.
METHODS: SHEE and SHEEC cell lines were used to extract NMPs. The quality of NMPs was monitored by Western blot analysis including DNA topoisomerase IIα, proliferation cell nuclear antigen (PCNA) and histone. NMPs of SHEE and SHEEC were analyzed by two-dimensional electrophoresis (2-DE), silver staining and PDQuest6.2 image analysis software. Three spots in which the differentially expressed NMPs were more obvious, were selected and analyzed with matrix-assisted laser desorption/ionization time of flying mass spectrometry (MALDI-TOF-MS) and database search.
RESULTS: Western blot analysis revealed that DNA topoisomerase IIα and PCNA were detected, and the majority of histones were deleted in NMPs of SHEE and SHEEC. After 2-DE image analysis by PDQuest6.2 software, the 2-DE maps were detected with an average of 106 ± 7.1 spots in SHEE and 132 ± 5.0 spots in SHEEC. Most of them were matched one another (r = 0.72), only 16 protein spots were found differing in intensity. Three NMPs including cytoskeletal tropomyosin, FK506-binding protein 6, similar to retinoblastoma binding protein 8 were preliminarily identified by MALDI- TOF-MS.
CONCLUSION: These differentially expressed NMPs may play an important role during malignant transformation from SHEE to SHEEC. Their separation and identification will contribute to searching for specific markers and probing into the pathogenesis of esophageal carcinoma.
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Affiliation(s)
- Xing-Dong Xiong
- Department of Biochemistry and Molecular Biology, Medical College, Shantou University, 22 Xinling Road, Shantou 515031, Guangdong Province, China
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Shen ZY, Xu LY, Chen MH, Li EM, Li JT, Wu XY, Zeng Y. Upregulated expression of Ezrin and invasive phenotype in malignantly transformed esophageal epithelial cells. World J Gastroenterol 2003; 9:1182-6. [PMID: 12800220 PMCID: PMC4611780 DOI: 10.3748/wjg.v9.i6.1182] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the correlation between ezrin expression and invasive phenotype formation in malignantly transformed esophageal epithelial cells.
METHODS: The experimental cell line employed in the present study was originated form the progressive induction of a human embryonic esophageal epithelial cell line (SHEE) by the E6E7 genes of human papillomavirus (HPV) type 18. The cells at the 35th passage after induction called SHEEIMM were in a state of immortalized phase and used as the control, while that of the 85th passage denominated as SHEEMT represented the status of cells that were malignantly transformed. The expression changes of ezrin and its mRNA in both cell passages were respectively analyzed by RT-PCR and Western blot. Invasive phenotype was assessed in vivo by inoculating these cells into the severe combined immunodeficient (SCID) mice via subcutaneous and intraperitoneal injection, and in vitro by inoculating them on the surface of the amnion membranes, which then was determined by light microscopy and scanning electron microscopy.
RESULTS: Upregulated expression of ezrin protein and its mRNA was observed in SHEEMT compared with that in SHEEIMM cells. The SHEEMT cells inoculated in SCID mice were observed forming tumor masses in both visceral organs and soft tissues in a period of 40 d with a special propensity to invading mesentery and pancreas, but did not exhibit hepatic metastases. Pathologically, these tumor cells harboring larger nucleus, nucleolus and less cytoplasm could infiltrate and destroy adjacent tissues. In the in vitro study, the inoculated SHEEMT cells could grow in cluster on the amniotic epithelial surface and intrude into the amniotic stroma. In contrast, unrestricted growth and invasiveness were not found in SHEEIMM cells in both in vivo and in vitro experiment.
CONCLUSION: The upregulated ezrin expression is one of the important factors that are possibly associated with the invasive phenotype formation in malignantly transformed esophageal epithelial cells.
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Affiliation(s)
- Zhong-Ying Shen
- Department of Pathology, Medical College of Shantou University, Shantou 515031, Guangdong Province, China.
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Shen ZY, Xu LY, Chen MH, Shen J, Cai WJ, Zeng Y. Progressive transformation of immortalized esophageal epithelial cells. World J Gastroenterol 2002; 8:976-81. [PMID: 12439909 PMCID: PMC4656402 DOI: 10.3748/wjg.v8.i6.976] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the progressive transformation of immortal cells of human fetal esophageal epithelium induced by human papillomavirus, and to examine biological criteria of sequential passage of cells, including cellular phenotype, proliferative rate, telomerase, chromosome and tumorigenicity.
METHODS: The SHEE cell series consisted of immortalized embryonic esophageal epithelium which was in malignant transformation when cultivated over sixty passages without co-carcinogens. Cells of the 10th, 31st, 60th and 85th passages were present in progressive development after being transfected with HPV. Cells were cultivated in a culture flask and 24-hole cultural plates. Progressive changes of morphology, cell growth, contact-inhibition, and anchorage-dependent growth characteristics were examined by phase contrast microscopy. The cell proliferation rate was assayed by flow cytometry. The modal number of chromosomes was analyzed. HPV18E6E7 was detected by Western blot methods and activities of telomerase were analyzed by TRAP. Tumorigenicity of cells was detected with soft agar plates cultivated and with tumor formation in SCID mice.
RESULTS: In morphological examination the 10th passage cells were in good differentiation, the 60th and 85th passages cells were in relatively poor differentiation, and the 31st passage cells had two distinct differentiations. The characteristics of the 85th and 60th passage cells were weakened at contact-inhibition and anchorage-dependent growth. Karyotypes of four stages of cells belonged to hyperdiploid or hypotriploid, and bimodal distribution of chromosomes appeared in the 31st and 60th passage cells. All of these characteristics combined with a increasing trend. The activities of telomerase were expressed in the latter three passages. Four fourths of SCID mice in the 85th passage cells and one fourth of SCID mice in the 60th passage cells developed tumors, but the cells in the 10th and 31st passage displayed no tumor formation.
CONCLUSION: In continual cultivation of fetal esophageal epithelial cells with transduction of HPV18E6E7, cells from the 10th to the 85th passage were changed gradually from preimmortal, immortal, precancerous to malignantly transformed stages. All of these changes were in a dynamic progressive process. The establishment of a continuous line of esophageal epithelium may provide a in vitro model of carcinogenesis induced by HPV.
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Affiliation(s)
- Zhong-Ying Shen
- Department of Tumor Pathology, Medical College of Shantou University, Guandong Province, China.
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Xiong XD, Xu LY, Shen ZY, Cai WJ, Luo JM, Han YL, Li EM. Identification of differentially expressed proteins between human esophageal immortalized and carcinomatous cell lines by two-dimensional electrophoresis and MALDI-TOF-mass spectrometry. World J Gastroenterol 2002; 8:777-81. [PMID: 12378614 PMCID: PMC4656560 DOI: 10.3748/wjg.v8.i5.777] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To identify the differentially expressed proteins between the human immortalized esophageal epithelial cell line (SHEE) and the malignant transformed esophageal carcinoma cell line (SHEEC), and to explore new ways for studying esophageal carcinoma associated genes.
METHODS: SHEE and SHEEC cell lines were used to separate differentially expressed proteins by two-dimensional electrophoresis. The silver-stained 2-D gels was scanned with EDAS290 digital camera system and analyzed with the PDQuest 6.2 Software. Six spots in which the differentially expressed protein was more obvious were selected and analyzed with matrix-assisted laser desorption/ionization time of flying mass spectrometry (MALDI-TOF-MS).
RESULTS: There were 107±4.58 and 115±9.91 protein spots observed in SHEE and SHEEC respectively, and the majority of these spots between the two cell lines matched each other (r = 0.772), only a few were expressed differentially. After analyzed by MALDI-TOF-MS and database search for the six differentially expressed proteins, One new protein as well as other five sequence-known proteins including RNPEP-like protein, human rRNA gene upstream sequence binding transcription factor, uracil DNA glycosylase, Annexin A2 and p300/CBP-associated factor were preliminarily identified.
CONCLUSION: These differentially expressed proteins might play an importance role during malignant transformation of SHEEC from SHEE. The identification of these proteins may serve as a new way for studying esophageal carcinoma associated genes.
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Affiliation(s)
- Xing-Dong Xiong
- Department of Biochemistry and Molecular Biology, Shantou University Medical College, Shantou 515031, Guangdong Province, China
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Syrjänen KJ. HPV infections and oesophageal cancer. J Clin Pathol 2002. [PMID: 12461047 DOI: 10.1136/jcp.55.10.721]] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/04/2022]
Abstract
The first reports suggesting an involvement of human papillomavirus (HPV) in the development of both benign and malignant squamous cell tumours of the oesophagus date back to 1982. Since then, a substantial amount of literature has accumulated on this subject, summarised in this review. To date, 239 oesophageal squamous cell papillomas have been analysed in 29 separate studies using different HPV detection methods, with HPV being detected in 51 (21.3%) cases. Many more squamous cell carcinomas have been analysed: of the 1485 squamous cell carcinomas analysed by in situ hybridisation, 22.9% were positive for HPV DNA, as were 15.2% of the 2020 cases tested by the polymerase chain reaction. In addition, evidence derived from large scale serological studies, animal experiments, and in vitro studies is discussed in the light of the highly variable geographical incidence rates of oesophageal carcinoma worldwide. It may be that the (multifactorial) aetiology of oesophageal cancer differs greatly between those geographical areas with a low risk and those with a high risk for this disease. Oncogenic HPV types seem to play an important causal role, particularly in high risk areas.
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Affiliation(s)
- K J Syrjänen
- Unità di Citoistopatologia, Laboratorio di Epidemiologia e Biostatistica, Istituto Superiore di Sanità, Viale Regina Elena 299, I-00161 Rome, Italy.
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Abstract
The first reports suggesting an involvement of human papillomavirus (HPV) in the development of both benign and malignant squamous cell tumours of the oesophagus date back to 1982. Since then, a substantial amount of literature has accumulated on this subject, summarised in this review. To date, 239 oesophageal squamous cell papillomas have been analysed in 29 separate studies using different HPV detection methods, with HPV being detected in 51 (21.3%) cases. Many more squamous cell carcinomas have been analysed: of the 1485 squamous cell carcinomas analysed by in situ hybridisation, 22.9% were positive for HPV DNA, as were 15.2% of the 2020 cases tested by the polymerase chain reaction. In addition, evidence derived from large scale serological studies, animal experiments, and in vitro studies is discussed in the light of the highly variable geographical incidence rates of oesophageal carcinoma worldwide. It may be that the (multifactorial) aetiology of oesophageal cancer differs greatly between those geographical areas with a low risk and those with a high risk for this disease. Oncogenic HPV types seem to play an important causal role, particularly in high risk areas.
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Affiliation(s)
- K J Syrjänen
- Unità di Citoistopatologia, Laboratorio di Epidemiologia e Biostatistica, Istituto Superiore di Sanità, Viale Regina Elena 299, I-00161 Rome, Italy.
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Shen ZY, Xu LY, Li EM, Cai WJ, Chen MH, Shen J, Zeng Y. Telomere and telomerase in the initial stage of immortalization of esophageal epithelial cell. World J Gastroenterol 2002; 8:357-62. [PMID: 11925625 PMCID: PMC4658384 DOI: 10.3748/wjg.v8.i2.357] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To search for the biomarker of cellular immortalization, the telomere length, telomerase activity and its subunits in cultured epithelial cells of human fetal esophagus in the process of immortalization.
METHODS: The transgenic cell line of human fetal esophageal epithelium (SHEE) was established with E6E7 genes of human papillomavirus (HPV) type 18 in our laboratory. Morphological phenotype of cultured SHEE cells from the 6th to 30th passages, was examined by phase contrast microscopy, the telomere length was assayed by Southern blot method, and the activity of telomerase was analyzed by telomeric repeat amplification protocol (TRAP). Expressions of subunits of telomerase, hTR and hTERT, were assessed by RT-PCR. DNA content in cell cycle was detected by flow cytometry. The cell apoptosis was examined by electron microscopy (EM) and TUNEL label.
RESULTS: SHEE cells from the 6th to 10th passages showed cellular proliferation with a good differentiation. From the 12th to the 16th passages, many senescent and apoptotic cells appeared, and the telomere length sharply shortened from 23 kb to 17 kb without expression of hTERT and telomerase activity. At the 20th passage, SHEE cells overcame the senescence and apoptosis and restored their proliferative activity with expression of telomerase and hTERT at low levels, but the telomere length shortened continuously to the lowest of 3 kb. After the 30th passage cells proliferation was restored by increment of cells at S and G2M phase in the cell cycle and telomerase activity expressed at high levels and with maintenance of telomere length.
CONCLUSION: At the early stage of SHEE cells, telomeres are shortened without expression of telomerase and hTERT causing cellular senescence and cell death. From the 20th to the 30th passages, the activation of telomerase and maintenance of telomere length show a progressive process for immortalization of esophageal epithelial cells. The expression of telomerase may constitute a biomarker for detection of immortalization of cells.
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Affiliation(s)
- Zhong-Ying Shen
- Department of Tumor Pathology, Medical College of Shantou University, 22 Xinling Road, Shantou 515031, Guandong Province, China.
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