Inhibition of CDH17 gene expression via RNA interference reduces proliferation and apoptosis of human MKN28 gastric cancer cells

Abstract

Gastric cancer is the fourth most common type of cancer and the second cause of cancer‑related mortalities worldwide despite the use of multimodal therapy. Cadherins are transmembrane glycoproteins that are involved in tumorigenesis. CDH17 has been found to be over‑expressed in gastric cancer and its overexpression was associated with lymph node metastasis and tumor‑node‑metastasis stage of the patients, yet the exact role and molecular mechanism of CDH17 in gastric cancer have not been determined. Using a lentiviral system as a delivery mediator of RNA interference, we found that inhibition of CDH17 can lead to reduce proliferation and increase apoptosis of gastric cancer cell line MKN28 in vitro and significantly diminish their tumorigenicity in vivo. Our results of the present study suggest that CDH17 may be a promising candidate for the therapeutic targeting of gastric cancer.

Figure 1

(A) CDH17 immunohistochemical staining was negative in normal gastric epithelium. (B) Non-neoplastic mucosa with intestinal metaplasia (H&E staining), (C) which revealed expression of CDH17. Immunohistochemical staining with CDH17 in (D) well-, (E) moderately and (F) poorly differentiated gastric cancer tissue (original magnification, ×400).

Figure 2

(A) Expression of CDH17 in MKN28 cells by immunofluorescence (magnification, ×1,000). (B) Fluorescence photomicrographs of gastric cancer cells infected by lenti-shCDH17. Images were captured 72 h after infection. (C) Quantitative PCR analysis showed the inhibition of CDH17 at the mRNA level in lenti-shCDH17 cells. (D) Decreased protein level of CDH17 as shown by western blotting. (Each bar is the mean ± SD. ^**P<0.01).

Figure 3

The effect of CDH17 downregulation on cell growth in vitro. MKN28 cells without any treatment and non-targeted RNAi vector-mediated lenti-shCDH17-neg cells were used as controls. (A) Cell migration. (B) Cell invasion. (C) Annexin V and propidium iodide double staining detected apoptosis. (D) Colony formation. (E) Cell proliferation. Knockdown of CDH17 caused a significant growth inhibition of gastric cancer cells as revealed by different assays. (F) Pro-caspase-3 and cleaved caspase-3 were examined by western blotting. ^*P<0.05; ^**P<0.01.

Figure 4

Knockdown of CDH17 inhibited tumor growth in a gastric cancer tumor model. (A) Images of tumors derived from MKN28, lenti-shCDH17-neg and lenti-shCDH17 cells in nude mice. (B) The changes in volume of the tumors of different groups. (C) Individual tumor weight at time of sacrifice. (D) Average body weight of nude mice measured at different time points. Data are presented as mean ± SD. ^**P<0.01.

Figure 5

(A) Pathological evaluation of tumor specimens from nude mice was determined by H&E staining (original magnification, ×400). Frozen sections of xenograft tissues were used to examine the expression levels of CDH17. The reduction of CDH17 in lenti-shCDH17 cells was detected by immunofluorescent staining. All the nuclei were stained with DAPI. The expression of CDH17 is shown in red (original magnification, x200). Increased level of TUNEL and cleaved caspase-3, and decreased expression of Ki-67 antigen detected by immunohistochemistry (original magnification, ×400). (B) Quantification of the ratio of positive staining in each tumor tissue region. Data are shown as mean ± SD. ^**P<0.01.