miR‑195‑5p regulates multi‑drug resistance of gastric cancer cells via targeting ZNF139

Abstract

Gastric cancer (GC) is one of the most common malignant tumors with a high mortality rate. Reversing the multi‑drug resistance (MDR) of GC offers the potential for significant enhancement of the effect of chemotherapy and improvement of prognosis. Aberrant microRNA expression can attribute to the pathogenesis of GC. However, the effects of microRNA (miR)‑195‑5p on the MDR of GC cells remains to be fully elucidated. In the present study, the effect of miR‑195‑5p in regulating the MDR of GC cells was investigated. Reverse transcription quantitative‑polymerase chain reaction was used to analyze the levels of miR‑195‑5p in GC cells. Western blot analysis was performed to analyze the protein levels of ZNF139, P‑gp, BCL‑2 and MRP1. The chemosensitivity of GC cells was determined by MTT. The results showed that the expression of miR‑195‑5p was decreased in poorly differentiated GC tissues with a higher chemosensitivity. The overexpression of miR‑195‑5p promoted the chemosensitivity of GC cells. Bioinformatics analysis indicated that Zing finger 139 (ZNF139) was a target of miR‑195‑5p. miR‑195‑5p negatively regulated the expression of ZNF139 by binding to its 3'‑untranslated region. The silencing of ZNF139 promoted the chemosensitivity of GC cells, and the downregulation of ZNF139 reversed the effect of miR‑195‑5p inhibitor on the chemosensitivity of GC cells. In conclusion, miR‑195‑5p regulated the MDR of GC cells via targeting ZNF139.

Figure 1.

Expression of miR-195-5p is decreased in poorly differentiated gastric cancer tissues with higher chemosensitivity. The levels of (A) miR-195-5p, and (B) p-gp, bcl-2 and mrp1 were analyzed by reverse transcription-quantitative polymerase chain reaction analysis. (C) Chemosensitivities of 5-FU and L-OHP were determined by MTT. **P<0.01, vs. well differentiated group. miR, microRNA; p-gp, p-glycoprotein; bcl-2, B-cell lymphoma 2; mrp1, multi-drug resistance-associated protein 1; 5-FU, 5-fluorouracil; L-OHP, oxaliplatin.

Figure 2.

miR-195-5p regulates the multi-drug resistance of gastric cancer cells. The levels of miR-195-5p were analyzed in MNK28 cells transfected with (A) miR-195-5p mimic and (B) miR-195-5p inhibitor. The protein levels of P-gp, BCL-2 and MRP1 were measured by western blot analysis in MNK28 cells transfected with (C) miR-195-5p mimic and (D) miR-195-5p inhibitor. The chemosensitivities of 5-FU and L-OHP were determined by MTT in MNK28 cells transfected with (E) miR-195-5p mimic and (F) miR-195-5p inhibitor. *P<0.05 and **P<0.01, vs. control group. miR, microRNA; P-gp, P-glycoprotein; BCL-2, B-cell lymphoma 2; MRP1, Multi-drug resistance-associated protein 1; 5-FU, 5-fluorouracil; L-OHP, oxaliplatin; NC, negative control; 195m, miR-195-5p mimic; NCI, negative control inhibitor; 195i, miR-195-5p inhibitor.

Figure 3.

miR-195-5p negatively regulates the expression of ZNF139 by binding to its 3′-UTR. (A) Bioinformatics database prediction of the binding site of miR-195-5p on the ZNF139 3′-UTR. (B) Nucleotides of the binding site were mutated. (C) Luciferase assay confirmed that miR-195-5p was able to bind to the 3′-UTR of ZNF139. The protein levels of ZNF139 were determined by western blot analysis in in MNK28 cells transfected with (D) miR-195-5p mimic and (E) miR-195-5p inhibitor. *P<0.05, vs. control group. miR, microRNA; ZNF139, Zing finger 139; 3′-UTR, 3′-untranslated region; WY, wild-type; Mut, mutated; NC, negative control; 195m, miR-195-5p mimic; NCI, negative control inhibitor; 195i, miR-195-5p inhibitor.

Figure 4.

Silencing of ZNF 139 promotes the chemosensitivity of gastric cancer cells. (A) Three pairs of siRNAs targeting ZNF139 were designed. (B) Protein levels of P-gp, BCL-2, MRP1 and ZNF139 were measured using western blot analysis in MNK28 cells transfected with siRNA-ZNF139. (C) Chemosensitivities of 5-FU and L-OHP were determined by MTT in MNK28 cells transfected with siRNA-ZNF139. *P<0.05 and **P<0.01, vs. control group. miR, microRNA; ZNF139, Zing finger 139; p-gp, p-glycoprotein; bcl-2, B-cell lymphoma 2; mrp1, multi-drug resistance-associated protein 1; 5-FU, 5-fluorouracil; L-OHP, oxaliplatin; CON, control; siRNA, small interfering RNA; NC, negative control.

Figure 5.

miR-195-5p regulates the multi-drug resistance of gastric cancer cells via targeting ZNF139. (A) Protein levels of P-gp, BCL-2, MRP1 and ZNF139 were measured in MNK28 cells using western blot analysis. (B) Chemosensitivities of 5-FU and L-OHP were determined by MTT in MNK28 cells transfected with siRNA-ZNF139 *P<0.05 and **P<0.01. miR, microRNA; ZNF139, Zing finger 139; P-gp, P-glycoprotein; BCL-2, B-cell lymphoma 2; MRP1, multi-drug resistance-associated protein 1; NCI, negative control inhibitor; 195i, miR-195-5p inhibitor; si, small interfering RNA; 5-FU, 5-fluorouracil; L-OHP, oxaliplatin.