Fig. 7: Cytotoxic activity of SaroL-1 against H1299 cells.

a Dose-dependent increase of cytotoxicity following the addition of purified SaroL-1 in a standard cell proliferation assay (MTT) demonstrating increment of cytotoxicity after 24 h of incubation compared to treatment with PBS. Cell viability is reduced by ~87% after stimulation with 1.36 µM SaroL-1. Data represent three independent experiments, n = 3. b Cell proliferation assay (MTT) of H1299 cells pre-treated with PPMP for 72 h before addition of increasing concentrations of purified SaroL-1. Cytotoxicity was remarkably reduced after 24 h of incubation with SaroL-1 in absence of the glycosphingolipid Gb3 at the plasma membrane of treated cells. Data represent three independent experiments, n = 3. c The soluble sugar PNPG inhibited SaroL-1 cytotoxicity. H1299 were incubated with increasing concentrations of SaroL-1 pre-treated with 10 mM PNPG. Cell proliferation assay (MTT) was used to assess SaroL-1´s cytotoxic activity after 24 h in comparison to the treatment with PBS. The results indicate that cell viability is preserved when SaroL-1 glycan-binding sites are saturated with soluble 10 mM PNPG. Data represent three independent experiments, n = 3. d H1299 cells suffer of acute cytotoxicity and membrane damage in presence of SaroL-1. A lactate dehydrogenase (LDH) assay revealed impairment of cell membrane integrity upon incubation with 0.27 µM and 1.36 µM SaroL-1 after 2 and 4 h. Data represent two independent experiments, n = 2. Differences to the control were analyzed for significance by using two-tailed unpaired t-test. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.