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26 February 2019 : Laboratory Research  

Cyclopamine Suppresses Human Esophageal Carcinoma Cell Growth by Inhibiting Glioma-Associated Oncogene Protein-1, a Marker of Human Esophageal Carcinoma Progression

Jing Yu1ABCDEF, Ruinuan Wu2B, Zhenyu Wang1C, Shuxian Chen1D, Suzuan Chen1E, Guanghua Guo1F, Zhaohui Liu3AG*

DOI: 10.12659/MSM.912858

Med Sci Monit 2019; 25:1518-1525

Abstract

BACKGROUND: Esophageal carcinoma is a common gastrointestinal tumor in humans. Cyclopamine, a Hedgehog (Hh)-pathway-specific inhibitor, is an effective chemotherapeutic drug for suppressing tumor cell differentiation, with unclear mechanisms. We investigated glioma-associated oncogene protein-1 (Gli-1) expression in human esophageal carcinoma tissue and the inhibition of cyclopamine on EC9706 esophageal carcinoma cell growth.

MATERIAL AND METHODS: Gli-1 in tumor tissue was measured by immunohistochemistry (IHC). EC9706 cells were treated with different concentrations of cyclopamine and incubated for different times. MTT method, flow cytometry, and Acridine orange/ethidium bromide (AO/EB) double-fluorescence staining were applied to detect cell proliferation and apoptosis. Western blot (WB) analysis was performed to assess Gli-1 expression.

RESULTS: Gli-1 was associated with patient age, gender, lymphatic metastasis, tumor recurrence, and stage, with significantly (P<0.05) positive correlations with age, lymphatic metastasis, tumor recurrence, and stage. At 12 h (F=214.57), 24 h (F=76.832), 48 h (F=236.90), and 72 h (F=164.55), the higher the concentration of cyclopamine, the higher the inhibition rate of suppressing EC9706 proliferation, and this effect was significant (P<0.05). The number of early-apoptosis cells increased as the concentration of cyclopamine increased. Morphology of EC9706 cells appeared as round with rough edges, karyopyknosis, and karyorrhexis. After 48 h, apoptosis rates of EC9706 cells treated with different concentrations of cyclopamine were (7.73±1.25)% at 2.5 μM, (13.37±1.42)% at 5.0 μM, (22.3±2.92)% at 10.0 μM, and (33.57±1.75)% at 20.0 μM, and the effect was dose-dependent. Gli-1 was obviously reduced after cyclopamine treatment and the effect was dose-dependent.

CONCLUSIONS: Gli-1 is highly expressed in human esophageal carcinoma, and could be a marker for use in assessing tumor stage and the deciding on treatment target.

Keywords: Esophageal Neoplasms, Glioma, Hedgehogs, Carcinoma, Squamous Cell, Cell Cycle, Disease Progression, Down-Regulation, Veratrum Alkaloids, Zinc Finger Protein GLI1

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Medical Science Monitor eISSN: 1643-3750
Medical Science Monitor eISSN: 1643-3750