Cellular Uptake and Intracellular Distribution of Phycocyanobilin in PC12 Cells: Spatial Association with Redox- and Mitochondrial-Related Regions

Background Phycocyanobilin (PCB) is a linear tetrapyrrolic chromophore derived from C-phycocyanin with reported antioxidant and neuroprotective properties. However, its cellular uptake and intracellular distribution in neuronal cells remain poorly characterized. This study aimed to characterize PCB internalization in PC12 cells and to examine its spatial association with DHE- and JC-1-positive intracellular regions. Methods PC12 cells were incubated with PCB at 25 or 100 μM for different time periods. Cellular uptake and accumulation were evaluated by confocal microscopy (CFM) and flow cytometry. Intracellular spatial association of PCB with DHE- and JC-1-positive regions was assessed by colocalization analysis using Pearson’s coefficient, Manders’ coefficients, Intensity Correlation Quotient (ICQ), and Costes randomization analysis. Results PCB was progressively internalized by PC12 cells and accumulated predominantly in cytoplasmic and perinuclear regions in a time- and concentration-dependent manner. CFM and flow cytometry showed strong concordance, with significant correlations for the percentage of PCB-positive cells (r = 0.8603, p = 0.0061) and fluorescence intensity (r = 0.9563, p = 0.0002). PCB showed substantial spatial overlap with DHE-positive regions, with high Manders’ coefficients (M1 = 0.99; M2 = 0.91) and positive ICQ values. Spatial association was also observed with JC-1-positive regions, although Pearson coefficients indicated a weaker linear intensity correlation than that observed with DHE. Conclusions PCB is efficiently internalized by PC12 cells and exhibits time- and concentration-dependent intracellular accumulation. Its spatial association with DHE- and JC-1-positive regions provides preliminary evidence linking intracellular PCB distribution to cellular regions relevant to redox and mitochondrial biology. Further studies using organelle-specific markers and functional assays are required to establish the mechanistic significance of these spatial associations.