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. 2018 Jan 8;8:93. doi: 10.1038/s41598-017-18551-z

Figure 2.

Figure 2

Knockdown of Sema4B inhibits proliferation and induces cell death. (A) qPCR results show that knockdown of endogenous Sema4B by three different shRNA sequences reduces the expression of this gene in U87-MG cells (data are presented as mean ± s.e.m., Mann–Whitney U test, n = 3). (B) XTT assay was used to evaluate U87-GM cell number at 1, 2, 3 and 4 days after plating. Each group of cells was treated with lentivirus expressing shRNA targeting Sema4B or control. The cells were plated for XTT assay 48 h after infection (data are presented as medians with range). P values were calculated with one-tailed Wilcoxon Signed Ranks Test (n = 5). (C–H) Effect of sh-cont and two sh-Sema4B were tested on U87-MG (C,E,G) or G55TL (D,F,H). (C,D) The same fields were monitored from 48–168 h (C) or 48–96 h (D) after infection (data are presented as mean ± s.e.m., n = 3). The p values were calculated with the Chi-square Fisher’s Exact test. (G,H) Glioma cell death was monitored using live/dead assay from 48 h up to 168 h (E) or up to 96 h (F). Data represent the means ± s.e.m. of three experiments (n = 3); p values were calculated with Chi-square Fisher’s Exact test. (G,H) Cell proliferation was monitored by adding a BrdU pulse 2 h before fixation. Proliferation was tested at 24–72 h (G) or 48–96 h (H). Percentage of BrdU-positive cells of the total DAPI-positive cells in each field is presented. Data represent the means ± s.e.m. of three experiments (n = 3); p values were calculated with Chi-square Fisher’s Exact test. *P < 0.05, **p < 0.001 ***p < 0.001.