Table 1.
Target primer of CHST15 gene.
Table 2.
Sequences of primer for RT-PCR.
Fig 1.
The knockdown effect of CHST15 siRNA on pancreatic cancer cell lines.
(A) Relative quantities of CHST15 mRNA in control siRNA treated and CHST15 siRNA treated PANC-1, MIA PaCa-2, Capan-1 and Capan-2 cells (n = 3 in each) were shown after normalization using GAPDH as an internal control. In each cell lines, there were significant differences about the reduction of CHST15 expression in samples treated with target-specific siRNA for 48 h. Asterisks (**, ***) p<0.01, P<0.001, respectively between control siRNA and CHST15 siRNA. (B) A WST-1 assay. The mean level of proliferation in control siRNA treated cells (black column) was expressed as a standard (1.0) and the data were shown relative to the standard as a proliferation index. Mean±SD (n = 3). Significant suppressive effects of CHST15 siRNA on cell proliferation were observed in PANC-1, MIA PaCa-2, Capan-1 and Capan-2. Asterisks (*, **) p<0.05, P<0.01, respectively between control siRNA and CHST15 siRNA. (C) Relative quantities of p21 mRNA in control siRNA treated and CHST15 siRNA treated PANC-1 cells (n = 3–5 in each). There was a slight increase and a significant increase of p21 mRNA in CHST15 siRNA treated PANC-1 cells and Capan-2 cells compared to control siRNA, respectively. Asterisk (*) p<0.05 between CHST15 siRNA and control siRNA.
Fig 2.
Effect of CHST15 siRNA on tumor growth in a xenograft model.
(A) The change of tumor volume. Data were expressed as mean ± SD (Untreated; n = 6, Control siRNA: n = 12, CHST15 siRNA; n = 10). Asterisks (*, ***) p<0.05, P<0.001, respectively between control siRNA and CHST15 siRNA. (B) Relative quantities of CHST15 mRNA in untreated, control siRNA treated and CHST15 siRNA treated xenografts at day 14. Data were expressed as mean±SD (Untreated; n = 6, Control siRNA: n = 12, CHST15 siRNA; n = 10).
Fig 3.
Effect of CHST15 siRNA on tumor histology in a xenograft model.
(A, B) Tumors in nude mice at day 19 were stained with HE (original magnification ×100). Representative pictures were shown. Widespread necrosis lesions were prominent in the CHST15 siRNA treated mice (A) compared to the control siRNA treated mice (B). (C, D, E, F, G) Immunohistochemical staining of CHST15 (brown, original magnifications ×100 for C, D, x400 for E, F, G) in xenografts at day 19. Representative pictures were shown. Although almost all tumor cells in xenograft were positive for CHST15 in control siRNA treated mice (D), strong positive signal was observed in the invasive front rather than the center of tumor. CHST15-highly positive tumor cells exhibited mesenchymal-like morphology (E) while CHST15-low positive cells exhibited cord-like morphology (F), In contrast, small number of remnant tumor cells was weakly positive for CHST15 in CHST15 siRNA treated mice (C).
Fig 4.
Effect of CS-E on PANC-1 tumor cell proliferation.
A WST-1 assay was performed using PANC-1 cell. The mean level of proliferation in PANC-1 cells without adding CS-E and HGF (white column) was expressed as a standard (1.0) and the data were shown relative to the standard as a proliferation index. The proliferation index in PANC-1 cells treated with low dose HGF (black column) did not show statistical difference in the culture system. In contrast, the proliferation index in HGF-treated PANC-1 cells significantly increased when adding CS-E (gray column). Data were expressed as mean±SD (n = 6 in each).