Galectin‑3 facilitates the proliferation and migration of nasopharyngeal carcinoma cells via activation of the ERK1/2 and Akt signaling pathways, and is positively correlated with the inflammatory state of nasopharyngeal carcinoma

Abstract

Nasopharyngeal carcinoma (NPC) is an epithelial carcinoma originating from the nasopharyngeal mucosal tissue and is highly prevalent in southeast Asia. Galectin‑3 (gal‑3) serves crucial roles in many cancers but its role in NPC remains to be elucidated. The aim of the present study was to investigate the role of gal‑3 in NPC. Immunohistochemistry and ELISA were used to determine the expression level of gal‑3 in patients with NPC or chronic rhinitis (CR). Gal‑3 short hairpin (sh)RNA was established to knockdown gal‑3 in 5‑8F and 6‑10B cells, allowing for the evaluation of the roles of gal‑3 in proliferation, migration and apoptosis in NPC cell lines. Immunohistochemistry staining of IL‑6 and IL‑8 was applied to access the inflammatory state of tumor tissues, and the correlation between the inflammatory state and gal‑3 was analyzed. The results demonstrated that gal‑3 was upregulated in patients with NPC compared with patients with CR. Knockdown of gal‑3 inhibited proliferation and migration in 5‑8F and 6‑10B cells, as well as promoted apoptosis in these cells. The expression levels of MMP‑9 and IL‑8 were also decreased in 5‑8F and 6‑10B cells after transfection with gal‑3 shRNA. A positive correlation was identified between the expression level of gal‑3 and the inflammatory state of NPC. The phosphorylation levels of ERK1/2 and Akt were downregulated after knockdown of gal‑3 in 5‑8F and 6‑10B cells. In conclusion, the expression level of gal‑3 was upregulated in patients with NPC and was positively correlated with the inflammatory state of NPC. The results suggested that gal‑3 promoted the proliferation and migration of 5‑8F and 6‑10B cells, while inhibiting the apoptosis of these cells. Moreover, activation of ERK1/2 and Akt may be the underlying mechanism of the effects of gal‑3 on NPC.

Keywords: galectin‑3; nasopharyngeal carcinoma; ERK1/2; Akt; inflammation.

Figure 1.

Expression levels of gal-3 are upregulated in patients with NPC and in NPC cell lines. (A) Representative images of immunochemistry staining of gal-3 in nasopharyngeal tissues and tumor tissues at ×400 magnification. (B) Concentrations of serum gal-3 in patients with NPC and patients with CR were determined via ELISA. ^#P<0.05. (C) mRNA expression levels of gal-3 in NP69, 5-8F and 6-10B cells were detected using reverse transcription-quantitative PCR and normalized to GAPDH. (D) Protein expression levels of gal-3 in NP69, 5-8F and 6-10B cells were detected via western blotting, using GAPDH as the loading control. **P<0.01, ***P<0.001 vs. NP69 cells. Gal-3, galectin-3; NPC, nasopharyngeal carcinoma; CR, chronic rhinitis.

Figure 2.

Knockdown of gal-3 inhibits the proliferation of 5-8F and 6-10B cells, while inducing their apoptosis. (A) Protein expression levels of gal-3 in 5-8F and 6-10B cells transfected with control shRNA or gal-3 shRNA were determined via western blotting, using GAPDH as the loading control. (B) mRNA expression levels of gal-3 in 5-8F and 6-10B cells transfected with control shRNA or gal-3 shRNA were determined via reverse transcription-quantitative PCR. (C) A MTT proliferation assay using was used to investigate the proliferation of 5-8F and 6-10B cells transfected with control shRNA or gal-3 shRNA. Representative images of Hoechst 33258 and EdU staining of (D) 5-8F and (E) 6-10B cells transfected with control shRNA or gal-3 shRNA to show their apoptosis and proliferation (magnification, ×400). ^†P<0.05, ^††P<0.01, ^†††P<0.001 vs. control shRNA. shRNA, short hairpin RNA; Gal-3, galectin-3; EdU, 5-Ethynyl-2′-deoxyuridine; O.D, optical density.

Figure 3.

Knockdown of gal-3 inhibits the migration of 5-8F and 6-10B cells. Representative images of the scratch assay demonstrating the mobility of (A) 5-8F and (B) 6-10B transfected with control shRNA or gal-3 shRNA (magnification, ×100). Representative images of a Transwell assay showing the capability of migration of (C) 5-8F and (D) 6-10B transfected with control shRNA or gal-3 shRNA (magnification, ×200). (E) mRNA expression levels of MMP-9 in 5-8F and 6-10B cells transfected with control shRNA or gal-3 shRNA were investigated via reverse transcription-quantitative PCR. ^†P<0.05, ^††P<0.01, ^†††P<0.001 vs. control shRNA. shRNA, short hairpin RNA; Gal-3, galectin-3.

Figure 4.

Gal-3 is associated with the inflammatory state of NPC. (A) Representative images of IHC staining gal-3, IL-6 and IL-8 in tumor tissues (magnification, ×400). The Spearman's correlation analyzation indicated that the expression level of gal-3 was positively correlated with the expression levels of IL-6 and IL-8 in tumor tissues. (B) mRNA expression levels of IL-8 in 5-8F and 6-10B cells transfected with control shRNA or gal-3 shRNA were determined using reverse transcription-quantitative PCR. ^††P<0.01, ^†††P<0.001 vs. control shRNA. shRNA, short hairpin RNA; Gal-3, galectin-3; IHC, immunohistochemistry.

Figure 5.

Knockdown of gal-3 suppresses the ERK1/2 and Akt signaling pathways. Phosphorylation levels of ERK1/2 and Akt in (A) 5-8F and (B) 6-10B cells transfected with control shRNA or gal-3 shRNA were detected via western blotting, using GAPDH as the loading control. ^†P<0.05, ^††P<0.01, ^†††P<0.001 vs. control shRNA. shRNA, short hairpin RNA; Gal-3, galectin-3; p-, phosphorylated.