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Figure 2 PDI Over-Expression Protects HL1 Cardiomyocytes From Apoptosis Under Hypoxic Conditions
(A) Endogenous protein disulfide isomerase (PDI) expression is induced under hypoxic conditions in mouse cardiomyocytes. HL1 cells were cultured under hypoxia conditions for 24 h and 48 h. The PDI expression was analyzed by Western blotting. (B) Schematic representation of PDI thioredoxin-like domains. The PDI variants have both cysteine-to-serine mutations in the N-terminal and/or C-terminal thioredoxin-like domains. Over-expression of wild-type PDI (PDI-wt), PDI-N, and PDI-C in HL1 stable individual clones were estimated by Western blot analysis. (C) Stable over-expression of PDI-wt but not of the N- and C-terminal mutants results in a significant protection from hypoxia-induced apoptosis. The HL1 cells were stable transfected with PDI-wt, PDI-N, PDI-C, or empty vector. Stable individual clones were incubated for the indicated periods in hypoxic conditions. The percentage of apoptotic cells was estimated by flow cytometric analysis. Values represent the mean ± SE of 5 independent experiments. *Significant differential apoptotic rate (Student t test, n = 5, p < 0.001) in normoxic and hypoxic cardiomyocytes at 24 h and 48 h. **Significant differential apoptotic rate (Student t test, n = 5, p < 0.001) in cells transfected with enzymatically inactive point mutants compared with the control in normoxic and hypoxic conditions at 24 h and 48 h.
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