Fig. 2: Characterization of SCCNVs.
From: Senescent cancer cell-derived nanovesicle as a personalized therapeutic cancer vaccine
a SA β-gal staining (green) of B16F10 cancer cells following senescence induction by treatment with various concentrations of doxorubicin for 4 days. Scale bars = 100 μm. b Optimization of the Dox concentration: the cell number relative to the initial cell number and nanovesicle production yield of B16F10 cancer cells after doxorubicin treatment at various concentrations for 4 days. c Optimization of the Dox concentration: antitumor efficacy of SCCNVs produced from cancer cells treated with various doxorubicin concentrations. n = 3–5. d Phosphatidylserine exposure on the B16F10 cancer cell surface after senescence induction with 0.1 μM doxorubicin, as evaluated by flow cytometric analysis. NT no treatment, SC senescence induction. n = 3. mRNA levels of (e) p16 and p21, (f) IFN-γ and TNF-α and (g) protein levels of IFN-γ and TNF-α in B16F10 cancer cells after senescence induction with 0.1 μM doxorubicin. NT no treatment, SC senescence induction. h Transmission electron microscopy image of SCCNVs. Scale bar = 250 nm. i Hydrodynamic diameter of SCCNVs. j Detection of the ovalbumin epitope presented on MHC class I expressed by E.G7-OVA cancer cells by FACS analysis. Following extrusion through nanosized pores, 15% of SCCNVs were positive for the OVA-MHC I complex. pH adjustment enabled the detached OVA in the supernatant to be reloaded on MHC I molecules, increasing the proportion of OVA-MHC I-positive SCCNVs to 96%. Relative (k) mRNA and (l) protein levels of IFN-γ and TNF-α in CCNVs and SCCNVs. m Hydrodynamic diameter of SCCNVs incubated in 10% (v/v) serum-containing buffer for various time periods, showing the colloidal stability of the SCCNVs. ns not significant. Data represent the mean ± SD. Statistical significance was calculated by Student’s t-tests (e, f, and k) or one-way analysis of variance (ANOVA) with Tukey’s multiple comparisons test (a, b, and m). *P < 0.05 versus NT or 0 μM doxorubicin in (a–f), versus CCNVs in i. †P < 0.05 versus 0.1 μM in a and b, ‡P < 0.05 versus 0.5 μM in b.
