The expression of microRNA-324-3p as a tumor suppressor in nasopharyngeal carcinoma and its clinical significance

Figures & data

Figure 1 Correlation between miR-324-3p expression and clinical features of NPC.

Notes: (A) The expression levels of miR-324-3p in 54 NPC tissues and 35 control tissues with no NPC (normal nasopharyngeal mucosa or chronic inflammation of nasopharyngeal mucosa) were detected by qRT-PCR. The relative expression level of miR-324-3p was calculated using 2-ΔΔCt method. miR-324-3p was downregulated in NPC tissues. (B) The correlation of miR-324-3p expression in NPC tissues and the overall survival of patients were analyzed by the Kaplan–Meier method.
Abbreviations: Cum, cumulative; NPC, nasopharyngeal carcinoma; qRT-PCR, quantitative reverse transcription-polymerase chain reaction.

Table 1 The relationship between miR-324-3p expression and the clinicopathologic characteristics in NPC patients

Variable	Cases (n)	Average expression of miR-324-3p	P-value
Sex			0.647
Male	36	2.48±0.45
Female	18	2.16±0.52
Age (years)			0.528
≥60	14	2.51±0.53
<60	40	2.17±0.48
Degree of differentiation			0.032
Well and moderately	5	3.47±0.71
Poorly	49	1.55±0.52
TNM stage			0.007
I–II	20	3.85±0.59
III–IV	34	1.02±0.47
Lymph node status			0.019
Metastasis	42	1.41±0.50
No metastasis	12	3.56±0.61

Abbreviation: NPC, nasopharyngeal carcinoma.

Figure 2 Effects of miR-324-3p-3p expression on migration and invasion of nasopharyngeal carcinoma cells.

Notes: (A) The results of qPCR showed that the expression of miR-324-3p in nasopharyngeal carcinoma 5–8F cells was significantly upregulated following 48 h transfection compared to the control. (B) The scratch healing experiments showed that 5–8F scratch healing ability was significantly decreased than the blank control group and negative control. (C) The data showed that the number of cells passing through the polycarbonate membrane was significantly lower in the miR-324-3p group than that in the blank control and the negative control group. (D) Western blot results showed that the expression of E-cadherin was increased, while the expression of vimentin was downregulated in nasopharyngeal carcinoma 5–8F cells. *P<0.05, compared to control cells.
Abbreviations: NC, negative control; qPCR, quantitative polymerase chain reaction.

Figure 3 Effect of miR-324-3p on cell viability and apoptosis.

Notes: (A) Cell proliferation rates were detected with MTT method. miR-324-3p significantly suppressed cell proliferation compared to control cells. (B) miR-324-3p mimic significantly induced cell apoptosis in 5–8F nasopharyngeal carcinoma cells. (C) The apoptosis results presented were obtained from Becton, Dickinson and Company (BD) flow cytometry software. Three independent experiments were performed. Data are shown as mean ± SD. *P<0.05 compared to control cells.
Abbreviation: NC, negative control.

Figure 4 GLI3 as the target gene of miR-324-3p-regulated NPC cell invasion.

Notes: (A) Luciferase results confirmed that miR-324-3p could directly regulate the expression of GLI3 gene. The luciferase activity in cells containing wild-type GLI3-3′-UTR was significantly inhibited, compared to cells with mutant form of GLI3-3′-UTR and control cells. (B) The GLI3 protein level in miR-324-3p mimic group was significantly lower than that in the control group. (C) The GLI3 protein level in miR-324-3p inhibitor group was significantly higher than that in the control group. (D) The knock-down efficiency of GLI3 genes was evaluated with qRT-PCR and Western blot. (E) The Transwell assay results showed that the number of transmembrane cells in 5–8F cells was significantly lower than that in the control group. Three independent experiments were performed. Data are shown as mean ± SD. *P<0.05 compared to the control cells.
Abbreviations: NC, negative control; NPC, nasopharyngeal carcinoma; qRT-PCR, quantitative reverse transcription-polymerase chain reaction; UTR, untranslated region.