Figure 1
(A) MiRNA expression in 3 GC tissues and adjacent normal tissues was analyzed by microarray analysis, (B) Quantification of microarray data showed that miR-147b was up-regulated in GC tissues. The data are expressed as mean multiple changes in miR-147b levels in GC tissues normalized to their corresponding controls.* P < 0.05, t test; (C) QPCR analysis showed that miR-147b was up-regulated in most GCs. The data shown are multiples of miR-147b expressions in GC cases normalized to paired adjacent normal tissues; (D) The expression of miR-147b in cultured GC cell lines and normal cells was analyzed by qPCR; (E) Wound healing analysis of BGC-823 and MGC-803 cells treated with miR-147b inhibitor or miR-147b NC.
Table 1
Expression of transfected miRNA-147b in normal cells and different gastric cancer cells detected by RT-PCR (x±s)
| Cell grouping | Number of samples | miR-147b | T value | Pvalue |
|---|---|---|---|---|
| Normal cell | 3 | 0.99±0.01 | ||
| SGC-7901 | 3 | 1.55±0.03 | 2.91 | <0.05 |
| BGC-823 | 3 | 1.74±0.12 | 2.91 | <0.05 |
| AGS | 3 | 1.47±0.02 | 2.35 | <0.05 |
| MKN-45 | 3 | 1.44±0.03 | 2.91 | <0.05 |
| MGC-803 | 3 | 1.8±0.04 | 2.91 | <0.05 |
[i] Note: miR-147bnc is an empty carrier
Figure 2
(A) After treatment with miR-147b inhibitor or miR-147b NC and staining with apoptosis markers (FITC annexin V and PI), the apoptosis of BGC-823 and MGC-803 cells was examined by flow cytometry. In the apoptosis diagram, FITC Annexin V + / PI + represents late apoptosis, FITC Annexin V + / pi - represents early apoptosis, and FITC Annexin V- / pi - represents normal living cells; (B) Clonogenic assay of BGC-823 and MGC-803 cells treated with miR-147b inhibitor or miR-147b NC. For all qPCR results, the data are expressed as mean ± SEM, and the error line represents the standard deviation obtained from three independent experiments; (C) Cell proliferation assay of BGC-823 and MGC-803 cells treated with miR-147b inhibitor of CCK-8 or miR-147b NC. (D) MiR-147b was up-regulated in GC cells.
Table 2
Effect of transfection of the miR-147b inhibitor on the number of clones of gastric cancer MGC-803 and BGC-823 cells
| Group | MGC-803 | BGC-823 |
|---|---|---|
| miR-147b nc | 638±15 | 632±17 |
| miR-147b inhibitor group | 464±11 | 448±15 |
| T value | 2.35 | 2.35 |
| P value | < 0.05 | < 0.05 |
[i] Note: miR-147b nc is an empty carrier
Table 3
Effect of transfection of the miR-147b inhibitor on the proliferation inhibition rate of gastric cancer cells by CCK-8 method (x±s)
| grouping Cell | Number samples of | miR-147bnc | miR-inhibitor 147b | T value | P value |
|---|---|---|---|---|---|
| MGC-803 | |||||
| 0h | 3 | 0.200±0.04 | 0.20±0.03 | 3.11 | >0.05 |
| 24h | 3 | 0.395±0.05 | 0.33±0.03 | 2.76 | >0.05 |
| 48h | 3 | 1.030±0.07 | 0.67±0.05 | 2.77 | <0.05 |
| 60h | 3 | 1.720±0.09 | 0.98±0.05 | 2.77 | <0.05 |
| 72h | 3 | 2.14±0.09 | 1.23±0.07 | 2.78 | <0.05 |
| BGC-823 | |||||
| 0 h | 3 | 0.20±0.04 | 0.20±0.03 | 3.07 | >0.05 |
| 24h | 3 | 0.49±0.05 | 0.33±0.03 | 2.78 | >0.05 |
| 48h | 3 | 1.13±0.07 | 0.62±0.05 | 2.18 | <0.05 |
| 60h | 3 | 1.56±0.09 | 0.85±0.05 | 2.77 | <0.05 |
| 72h | 3 | 1.84±0.1 | 1.02±0.07 | 2.78 | <0.05 |
[i] Note: miR-147b nc is an empty carrier
Figure 3
(A) Double luciferase reporter vector of CPEB2 wild type and mutant, (B) Expression of miR-147b inhibitor accelerates CPEB2 3 ‘- UTR activity, (C) Expression of CPEB2 in MGC-803 cells of different groups.
Figure 4
(A)Expression of CPEB2 mRNA in GC tissues and adjacent normal tissues, (B) Nonparametric test results of the relationship between CPEB2 expression and clinicopathological factors, (C)Overall survival and disease-free survival in GC patients with lower CPEB2 expression, (D) immunohistochemical analysis results; (E) Results of immunoblotting.
Table 4
Correlation between clinicopathological features and miR-147 expression in gastric cancer
| Variance | Number of samples | miR-147b Low(n%) | expression High(n%) | χ2 | P value |
|---|---|---|---|---|---|
| Age | |||||
| <60 | 22 | 4 (18.1%) | 18 (81.9%) | 0.081 | P >0.05 |
| ≥ 60 | 28 | 6 (21.4%) | 22 (78.6%) | ||
| Gender | |||||
| male | 29 | 7 (24.1%) | 22 (75.9%) | 0.176 | P >0.05 |
| female | 21 | 3 (14.3%) | 17 (85.7%) | ||
| Tumor size | |||||
| <4 cm | 33 | 6 (18.2%) | 27 (81.8%) | 0.006 | P >0.05 |
| ≥4 cm | 17 | 4 (23.5%) | 13 (76.5%) | ||
| TNM Stage | |||||
| I-II | 20 | 7 (35%) | 13 (65%) | 3.755 | P =0.048 |
| III-IV | 30 | 3 (10%) | 27 (90%) | ||
| Lymph node metastasis | |||||
| No | 23 | 8 (34.8%) | 15 (65.2%) | 4.232 | P =0.040 |
| Yes | 27 | 2 (7.4%) | 25 (92.6%) |