Low Molecular Weight Fucoidan Alleviates Cardiac Dysfunction in Diabetic Goto-Kakizaki Rats by Reducing Oxidative Stress and Cardiomyocyte Apoptosis

Figure 6

LMWF directly inhibited PKCβ-mediated ROS production in H9c2 cells induced by high glucose. ((a), (b)) H9c2 cells were incubated in normal glucose (NG, 5.5 mM D-glucose) and treated with high glucose (HG, 25 mM D-glucose) without or with LMWF (20 μg/mL) treatment for 24 h. D-mannitol (20 mM) was used as an osmotic control. Then cells were incubated with 20 μM H₂DCF-DA at 37°C for 45 min. After washing with PBS, fluorescence intensity was examined by fluorescence microscope (excitation wavelength = 485 nm, exposure time = 10 ms). The micrographs were taken by both light and fluorescence microscopes in the same fields. (c) H9c2 cells were seeded in 96-well microplates and subjected to the same treatment as above to detect ROS production. H₂O₂ was used as a positive control. (d) H9c2 cells were seeded in 96-well plates and subjected to the same treatment as above, after 72 h, cell viability was determined by MTT assay. (e) H9c2 cells were seeded in 6-well plates and were treated with HG (25 mM D-glucose) without or with LMWF (10, 20 μg/mL) treatment for 48 h. Then cells were harvested and protein lysates were subjected to Western blot analysis of the expression of PKCβ1 and PKCβ2 relative to the expression of α-actin. The densitometry value of PKC was normalized to that of α-actin and statistical analysis was performed. Data are presented as the mean ± SEM. versus control or D-mannitol group, , versus HG group.