Additional file 4 of Chitinase-3-like protein 1 depletion in glioma cells alters tumor microenvironment and normalizes neovasculature in human glioma xenografts

Supplementary Material 4: Figure 4. a. Gating strategy for the flow cytometric sort of U251 transfected with pCMV-Cas9-GFP plasmid. Percent of positive GFP signal is presented. b-c. Validation of CHI3L1 knock-out in selected sgCTR and CHI3L1 KO clones. b. RT-qPCR analysis of CHI3L1 expression. * P ≤ 0.05. c. ELISA of cell culture supernatants of sgCTR and CHI3L1 KO clones. ** P ≤ 0.01. d. MTT metabolic activity assay for selected sgCTR and CHI3L1 KO clones. e Gene expression analysis for MMP2 and IL-1β in CHI3L1 KO cells. Black line represents gene expression for sgCTR. * P ≤ 0.05, ** P ≤ 0.01. f. Matrigel invasion assay for U251 sgCTR and CHI3L1 KO cells. Black bars represent baseline invasiveness of glioma cells; grey and red dots/bars represent invasiveness of sgCTR and CHI3L1 KO cells when co-cultured with HM-SV40 human microglial cells; mean ± SD is presented. * P ≤ 0.05.