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. 2023 Mar 11;30(5):1366–1381. doi: 10.1038/s41418-023-01143-y

Fig. 6. Differential cell death activity of GSDMB isoforms in breast cancer cells.

Fig. 6

SKBR3 cell line was transiently transfected with GSDMB constructs during 48 h. A Cytotoxicity was measured by lactate dehydrogenase (LDH) assay. Values represent means ± SEMs of 4 independent experiments. Differences between control condition (empty vector) and each condition were tested by two-tailed unpaired t-test: *p < 0,05 and **>p < 0,01. GSDMD-NT was used as a positive control. B Immunofluorescence and confocal microscopy analysis in SKBR3 cells transiently transfected with the indicated GSDMB-NT constructs. GSDMB-NT (green), colocalizes with mitochondrial marker TOM20 (red). pLVX (empty vector) was used as negative control. C Lactate dehydrogenase (LDH) assay after 16 h of co-culture with NK-92. SKBR3 cells expressing GSDMB isoforms 2 and 3 (1-416Δ6,7, 1-416, respectively) were co-cultured with NK-92 cells. Values represent means ± SEMs of seven independent experiments. Differences between control condition (empty vector co-culture with NK-92) and each condition were tested by two-tailed unpaired t-test: **p < 0,01. D Immunoblotting of GSDMB cleavage by GZMA in SKBR3 cells expressing GSDMB isoforms (2 and 3) cocultured with NK-92 cells. Immunoblot antibodies for GSDMB cleavage detection: anti-GSDMB-NT (Sigma, HPA023925) and anti-GSDMB-CT antibody Ab-GB [39]. GAPDH was used as a loading control.