P L Reshma1, Neethu S Sainu1, Anil K Mathew2, K G Raghu3. 1. Agroprocessing and Natural Products Division, CSIR - National Institute for Interdisciplinary Science and Technology, Thiruvananthapuram, 695019, Kerala, India. 2. Biotechnology Division, CSIR - National Institute for Interdisciplinary Science and Technology, Thiruvananthapuram, 695019, Kerala, India. 3. Agroprocessing and Natural Products Division, CSIR - National Institute for Interdisciplinary Science and Technology, Thiruvananthapuram, 695019, Kerala, India. Electronic address: raghukgopal2009@gmail.com.
Abstract
AIMS: The present study investigates the protective effect of partially characterized Tribulus terrestris L. fruit methanol extract against mitochondrial dysfunction in cell based (H9c2) myocardial ischemia model. MAIN METHODS: To induce ischemia, the cells were maintained in an ischemic buffer (composition in mM -137 NaCl, 12 KCl, 0.5 MgCl2, 0.9 CaCl2, 20 HEPES, 20 2-deoxy-d-glucose, pH-6.2) at 37°C with 0.1% O2, 5% CO2, and 95% N2 in a hypoxia incubator for 1h. Cells were pretreated with various concentrations of T. terrestris L. fruit methanol extract (10 and 25μg/ml) and Cyclosporin A (1μM) for 24h prior to the induction of ischemia. KEY FINDINGS: Different parameters like lactate dehydrogenase release, total antioxidant capacity, glutathione content and antioxidant enzymes were investigated. Studies were conducted on mitochondria by analyzing alterations in mitochondrial membrane potential, integrity, and dynamics (fission and fusion proteins - Mfn1, Mfn2, OPA1, Drp1 and Fis1). Various biochemical processes in mitochondria like activity of electron transport chain (ETC) complexes, oxygen consumption and ATP production was measured. Ischemia for 1h caused a significant (p≤0.05) increase in LDH leakage, decrease in antioxidant activity and caused mitochondrial dysfunction. T. terrestris L. fruit methanol extract pretreatment was found effective in safeguarding mitochondria via its antioxidant potential, mediated through various bioactives. HPLC of T. terrestris L. fruit methanol extract revealed the presence of ferulic acid, phloridzin and diosgenin. SIGNIFICANCE: T. terrestris L. fruit ameliorate ischemic insult in H9c2 cells by safeguarding mitochondrial function. This validates the use of T. terrestris L. against heart disorders.
AIMS: The present study investigates the protective effect of partially characterized Tribulus terrestris L. fruit methanol extract against mitochondrial dysfunction in cell based (H9c2) myocardial ischemia model. MAIN METHODS: To induce ischemia, the cells were maintained in an ischemic buffer (composition in mM -137 NaCl, 12 KCl, 0.5 MgCl2, 0.9 CaCl2, 20 HEPES, 20 2-deoxy-d-glucose, pH-6.2) at 37°C with 0.1% O2, 5% CO2, and 95% N2 in a hypoxia incubator for 1h. Cells were pretreated with various concentrations of T. terrestris L. fruit methanol extract (10 and 25μg/ml) and Cyclosporin A (1μM) for 24h prior to the induction of ischemia. KEY FINDINGS: Different parameters like lactate dehydrogenase release, total antioxidant capacity, glutathione content and antioxidant enzymes were investigated. Studies were conducted on mitochondria by analyzing alterations in mitochondrial membrane potential, integrity, and dynamics (fission and fusion proteins - Mfn1, Mfn2, OPA1, Drp1 and Fis1). Various biochemical processes in mitochondria like activity of electron transport chain (ETC) complexes, oxygen consumption and ATP production was measured. Ischemia for 1h caused a significant (p≤0.05) increase in LDH leakage, decrease in antioxidant activity and caused mitochondrial dysfunction. T. terrestris L. fruit methanol extract pretreatment was found effective in safeguarding mitochondria via its antioxidant potential, mediated through various bioactives. HPLC of T. terrestris L. fruit methanol extract revealed the presence of ferulic acid, phloridzin and diosgenin. SIGNIFICANCE: T. terrestris L. fruit ameliorate ischemic insult in H9c2 cells by safeguarding mitochondrial function. This validates the use of T. terrestris L. against heart disorders.
Keywords:
Antioxidant enzymes; Electron transport chain; H9c2; Ischemia; Mitochondria; Mitochondrial fusion and fission; Oxidative phosphorylation; Oxidative stress; Tribulus terrestris L.