Suppression of colorectal cancer metastasis by nigericin through inhibition of epithelial-mesenchymal transition.
OBJECTIVE
To evaluate the effect of nigericin on colorectal cancer and to explore its possible mechanism.


METHODS
The human colorectal cancer (CRC) cell lines HT29 and SW480 were treated with nigericin or oxaliplatin under the conditions specified. Cell viability assay and invasion and metastasis assay were performed to evaluate the effect of nigericin on CRC cells. Sphere-forming assay and soft agar colony-forming assay were implemented to assess the action of nigericin on the cancer stem cell properties of CRC cells undergone epithelial-mesenchymal transition (EMT).


RESULTS
Compared with oxaliplatin, nigericin showed more toxicity for the HT29 cell line (IC50, 12.92 ± 0.25 μmol vs 37.68 ± 0.34 μmol). A similar result was also obtained with the SW116 cell line (IC50, 15.86 ± 0.18 μmol vs 41.02 ± 0.23 μmol). A Boyden chamber assay indicated that a significant decrease in the number of HT29 cells migrating through polyvinylidene fluoride membrane was observed in the nigericin-treated group, relative to the vehicle-treated group [11 ± 2 cells per high-power field (HPF) vs 19.33 ± 1.52 cells per HPF, P < 0.05]. Compared to the control group, the numbers of HT29 cells invading through the Matrigel-coated membrane also decreased in the nigericin-treated group (6.66 ± 1.52 cells per HPF vs 14.66 ± 1.52 cells per HPF, P < 0.05). Nigericin also reduced the proportion of CD133+ cells from 83.57% to 63.93%, relative to the control group (P < 0.05). Nigericin decreased the number of spheres relative to the control group (0.14 ± 0.01 vs 0.35 ± 0.01, P < 0.05), while oxaliplatin increased the number of spheres relative to the control group (0.75 ± 0.02 vs 0.35 ± 0.01; P < 0.05). Nigericin also showed a decreased ability to form colonies under anchorage-independent conditions in a standard soft agar assay after 14 d in culture, relative to the control group (1.66 ± 0.57 vs 7 ± 1.15, P < 0.05), whereas the colony numbers were higher in the oxaliplatin group relative to the vehicle-treated controls (14.33 ± 0.57 vs 7 ± 1.15, P < 0.05). We further detected the expression of E-cadherin and vimentin in cells treated with nigericin and oxaliplatin. The results showed that HT29 cells treated with nigericin induced an increase in E-cadherin expression and a decrease in the vimentin expression relative to vehicle controls. In contrast, oxaliplatin downregulated the expression of E-cadherin and upregulated the expression of vimentin in HT29 cells relative to vehicle controls.


CONCLUSIONS
This study demonstrated that nigericin could partly reverse the EMT process during cell invasion and metastasis.
Department of General Surgery, Ruijin Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai Minimally Invasive Surgery Center, Shanghai 200025, China.
Hou-Min
HM
Zhou
Tao-Tao
TT
Dong
Lin-Lin
LL
Wang
Bo
B
Feng
Hong-Chao
HC
Zhao
Xiu-Ke
XK
Fan
Min-Hua
MH
Zheng
D000071916:AC133 Antigen; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D015703:Antigens, CD / Q000378:metabolism; D015820:Cadherins / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement; D002470:Cell Survival; D015179:Colorectal Neoplasms / Q000188:drug therapy*; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects*; D005434:Flow Cytometry / Q000379:methods; D015972:Gene Expression Regulation, Neoplastic; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D009361:Neoplasm Invasiveness; D009362:Neoplasm Metastasis; D014411:Neoplastic Stem Cells / Q000166:cytology; D009550:Nigericin / Q000494:pharmacology*; D009944:Organoplatinum Compounds / Q000494:pharmacology; D000077150:Oxaliplatin; D010455:Peptides / Q000378:metabolism; D014746:Vimentin / Q000378:metabolism
D000071916:AC133 Antigen; D000900:Anti-Bacterial Agents; D015703:Antigens, CD; D015820:Cadherins; D006023:Glycoproteins; D009944:Organoplatinum Compounds; C000606023:PROM1 protein, human; D010455:Peptides; D014746:Vimentin; D000077150:Oxaliplatin; D009550:Nigericin