Fig. 5. ManN specifically induces expression of unfolded protein response (UPR) responsive proteins.

a BCECs were cultured in growth media (GM) until ~80% confluency. Media were changed to growth factor-free media containing 10% BCS in the presence or absence of 40 or 400 μM of ManN or mannose for various times. At the end of each incubation, cell lysates were collected, proteins were separated on 4–12% Bis-Tris gel for western blot analysis. b Cells were treated with various concentrations of ManN, mannose, 5 ng/ml VEGF, or a combination of ManN and VEGF for 24 h. Cell lysates were separated on NuPAGE 3–8% Tris-Acetate gel for western blot analysis. c 4-PBA, but not TUDCA, could effectively block the induction of CHOP in BCECs, accompanied by a restoration of expression of transcription factor ATF-6 upon 400 μM ManN treatment. BCECs were pre-treated with 2 mM 4-PBA or 500 μM TUDCA, two chemical chaperons. Sixteen hours later, cells were switched to growth factor-free media for 4 h in the presence of ManN. GM: growth media. d 4-PBA significantly blocked the bell-shaped effects of ManN on BCEC proliferation. Pre-treatment of cells with 1 mM 4-PBA for 8 h abrogated additive effects of 40 μM ManN and 5 ng/ml VEGF and protected cells from toxic effect induced by 2 mM ManN. n = 3 independent samples. For each study, a representative experiment is shown from two to three independent studies. Data are means +/− SD, asterisks indicate a significant difference compared with control. Statistical analysis was done by two-tailed, two-sample unequal variance t test. *p < 0.05, **p < 0.01. Data are provided as a Source data file.