A phenylpropenoic acid glucoside (PPAG) of Aspalathus linearis protects H9c2 cardiomyocytes against hyperglycemia-induced cell apoptosis

Abstract

Hyperglycemia is responsible for altered myocardial substrate metabolism and subsequent apoptosis in diabetic patients. Adult cardiomyocytes have a limited capacity to regenerate after an injury. Thus, protecting the myocardium against chronic hyperglycemia is crucial for cell survival. Phenylpyruvic acid-2-O-β-D-glucoside (PPAG) is one of the major polyphenolic compounds found in Aspalathus linearis. PPAG has been shown to protect the pancreatic β-cells from animals with elevated blood glucose levels against cell apoptosis. Therefore, this study investigates the potential anti-apoptotic effect of PPAG against hyperglycemia-induced cardiac injury. H9c2 cardiomyocytes exposed to either normal (5.5mM) or high (33 mM) glucose for 48hrs were treated with PPAG (1 µM) as well as a combination of metformin and PPAG at 1 µM for 12hrs. The efficacy of PPAG to reverse altered cardiac energy metabolism was investigated by measuring the uptake and oxidation of fatty acids. Mitochondrial membrane depolarization was assessed by measuring JC-1 fluorescence while apoptotic cell death was determined by annexin V/propidium iodide staining in addition to caspase 3/7 activity. Results showed that high glucose exposure resulted in an increased fatty acid uptake and oxidation (35 ± 4%, p < 0.0001 and 39 ± 7%, p < 0.0001, respectively) when compared to the normal glucose control. In addition, an increase in membrane depolarization (38 ± 4%, p < 0.0001), annexin V/propidium iodide (36 ± 2%, p < 0.0001/37 ± 5%, p < 0.0001) and caspase 3/7 activity (32 ± 6%, p < 0.001) was observed. PPAG effectively reduced membrane depolarization (10.8 ± 1%, p < 0.0003), while treatment with a combination of metformin and PPAG had more effect in reducing annexin V/propidium iodide (14 ± 3%, p < 0.0005/12 ± 2%, p < 0.001) and caspase 3/7 activity (8 ± 1%, p < 0.007). Hence, suggesting that a combination of metformin and PPAG might be an effective treatment to protect the myocardium against high glucose-induced cell apoptosis.