FIG. 4.

Reperfusion with acidic media (“Rep [pH 6.4]”) to prevent rapid intracellular pH normalization and reduce ischemia–reperfusion injury. (A) Schematic of the experimental protocol for the comparison groups. (B) Measurement of intracellular pH using pHrodo dye. Higher fluorescence signal indicates a lower intracellular pH. Data are from two independent experiments. Constructs were analyzed for (C, D) cell death as measured by LDH and AK release, (E) cell viability as measured using RealTime-Glo assay, and (F) mitochondrial membrane permeability as measured by JC-1 dye, where increased ratio of emission at 528 and 590 nm is correlated with higher permeability. Data are from four independent experiments and are shown as individual data points with mean ± SD. Statistical analysis was done using ANOVA with post-hoc Tukey's HSD, ^#Indicates statistical significance compared to all other groups, *Indicates significant difference between groups, p < 0.05. (G) Western blot analysis of apoptosis by cleaved to total caspase 3 ratio. Data are from three independent experiments, mean ± SD. *Indicates significant difference between groups as determined by unpaired t-test, p < 0.05. ns, not significant. Color images are available online.