cytochrome-c-t and asiatic-acid

Asiatic acid, one of the triterpenoid components isolated from Centella asiatica, has received increasing attention due to a wide variety of biological activities. To date, little is known about its mechanisms of action. Here we examined the cytotoxic effect of asiatic acid on HepG2 cells and elucidated some of the underlying mechanisms. Asiatic acid induced rapid cell death, as well as mitochondrial membrane potential (MMP) dissipation, ATP depletion and cytochrome c release from mitochondria to the cytosol in HepG2 cells. In mitochondria isolated from mouse liver, asiatic acid treatment significantly stimulated the succinate-supported state 4 respiration rate, dissipated the MMP, increased Ca(2+) release from Ca(2+)-loaded mitochondria, decreased ATP content and promoted cytochrome c release, indicating the uncoupling effect of asiatic acid. Hydrogen peroxide (H2O2) produced by succinate-supported mitochondrial respiration was also significantly inhibited by asiatic acid. In addition, asiatic acid inhibited Ca(2+)-induced mitochondrial swelling but did not induce mitochondrial swelling in hyposmotic potassium acetate medium which suggested that asiatic acid may not act as a protonophoric uncoupler. Inhibition of uncoupling proteins (UCPs) or blockade of adenine nucleotide transporter (ANT) attenuated the effect of asiatic acid on MMP dissipation, Ca(2+) release, mitochondrial respiration and HepG2 cell death. When combined inhibition of UCPs and ANT, asiatic acid-mediated uncoupling effect was noticeably alleviated. These results suggested that both UCPs and ANT partially contribute to the uncoupling properties of asiatic acid. In conclusion, asiatic acid is a novel mitochondrial uncoupler and this property is potentially involved in its toxicity on HepG2 cells.

Topics: Adenosine Triphosphate; Animals; Calcium; Cell Death; Cell Respiration; Cytochromes c; Hep G2 Cells; Humans; Membrane Potential, Mitochondrial; Mice; Mitochondria, Liver; Mitochondrial Uncoupling Proteins; Pentacyclic Triterpenes; Reactive Oxygen Species

Asiatic acid (AA) is a pleiotropic neuroprotective agent that has been shown to attenuate infarct volume in mouse and rat models of focal ischemia and has a long clinically relevant therapeutic time-window. Because in a future trial AA would be administered with tissue-plasminogen activator (t-PA), the only approved acute stroke therapy, we sought to determine the effect of AA when co-administered with t-PA in a rat focal embolic stroke model. Male rats were treated with AA (75 mg/kg) alone, low-dose t-PA (2.5 mg/kg) alone, or a combination of AA and low-dose t-PA at 3 h after inducing embolic stroke. AA significantly reduced infarct volume whereas low-dose t-PA alone did not reduce infarct volume compared with vehicle. Significantly, combination treatment further enhanced reduction of infarct volume versus AA alone. Treatment with AA reduced cytochrome c (CytoC) and apoptosis-inducing factor (AIF) release from brain mitochondria after ischemia. AA was also neuroprotective against L-glutamate-induced toxicity in primary cortical neurons. In summary, combination treatment with AA and low-dose t-PA at 3 h after embolic stroke reduces infarct volume, improves neurological outcome, and provides neuroprotection. The neuroprotective effects of AA were partially associated with reduction of AIF and CytoC release.

Topics: Animals; Apoptosis Inducing Factor; Cytochromes c; Disease Models, Animal; Infarction, Middle Cerebral Artery; Male; Mitochondria; Neurons; Neuroprotective Agents; Pentacyclic Triterpenes; Rats, Sprague-Dawley; Rats, Wistar

Asiatic acid, a triterpenoid derivative from Centella asiatica, has shown biological effects such as antioxidant, antiinflammatory, and protection against glutamate- or beta-amyloid-induced neurotoxicity. We investigated the neuroprotective effect of asiatic acid in a mouse model of permanent cerebral ischemia. Various doses of asiatic acid (30, 75, or 165 mg/kg) were administered orally at 1 hr pre- and 3, 10, and 20 hr postischemia, and infarct volume and behavioral deficits were evaluated at day 1 or 7 postischemia. IgG (blood-brain barrier integrity) and cytochrome c (apoptosis) immunostaining was carried out at 24 hr postischemia. The effect of asiatic acid on stress-induced cytochrome c release was examined in isolated mitochondrial fractions. Furthermore, its effects on cell viability and mitochondrial membrane potential were studied in HT-22 cells exposed to oxygen-glucose deprivation. Asiatic acid significantly reduced the infarct volume by 60% at day 1 and by 26% at day 7 postischemia and improved neurological outcome at 24 hr postischemia. Our studies also showed that the neuroprotective properties of asiatic acid might be mediated in part through decreased blood-brain barrier permeability and reduction in mitochondrial injury. The present study suggests that asiatic acid may be useful in the treatment of cerebral ischemia.

Topics: Animals; Brain; Brain Ischemia; Cell Hypoxia; Cell Line; Cell Survival; Cytochromes c; Disease Models, Animal; Glucose; Immunoglobulin G; Infarction, Middle Cerebral Artery; Male; Membrane Potential, Mitochondrial; Mice; Mice, Inbred C57BL; Mitochondria; Neuroprotective Agents; Pentacyclic Triterpenes; Severity of Illness Index; Time Factors; Treatment Outcome; Triterpenes

Cancer is one of the leading causes of death in the world. The triterpenoid compound asiatic acid derived from the tropical medicinal plant Centella asiatica displays cytotoxic activity on fibroblast cells and several other kinds of cells. The present work studies asiatic acid-mediated growth inhibition of cancer cells and the underlying mechanism. Asiatic acid markedly inhibited cancer cell proliferation. Apoptosis of SW480 human colon cancer cells was induced by asiatic acid as shown by flow cytometry, DNA fragmentation and nuclear chromatin condensation experiments. Through increasing mitochondrial membrane permeability and cytochrome c release from mitochondria into cytosol, asiatic acid induced caspase-9 activity, which further activated caspase-3 and poly(ADP-ribose) polymerase cleavage resulting in irreversible apoptotic death in the tumor cells. Taken together, these results suggest that mitochondrial death apoptosis cascade plays very important roles in asiatic acid-induced cancer apoptosis.

Topics: Animals; Antineoplastic Agents, Phytogenic; Apoptosis; Blotting, Western; Caspase 3; Caspase 9; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Centella; Chromatin; Colonic Neoplasms; Colorectal Neoplasms; Cytochromes c; DNA Fragmentation; Flow Cytometry; Humans; Membrane Potential, Mitochondrial; Mice; Mice, Inbred BALB C; Microscopy, Confocal; Microscopy, Fluorescence; Mitochondria; Molecular Structure; Pentacyclic Triterpenes; Poly Adenosine Diphosphate Ribose; Stomach Neoplasms; Triterpenes

Asiatic acid (AA) is one of the triterpenoid components of Terminalia catappa L., which has antioxidative, anti-inflammatory and hepatoprotective activity. This research focused on the mitochondrial protection of AA against acute liver injury induced by lipopolysaccharide (LPS) and D-galactosamine (D-GalN) in mice. It was found that pretreatment with 25, 50 or 100 mg kg(-1) AA significantly blocked the LPS + D-GalN-induced increase in both serum aspartate aminotransferase (sAST) and serum alanine aminotransferase (sALT) levels, which was confirmed by ultrastructural observation under an electron microscope, showing improved nuclear condensation, ameliorated mitochondrion proliferation and less lipid deposition. Meanwhile, different doses of AA could decrease both the transcription and the translation level of voltage-dependent anion channels (VDACs), the most important mitochondrial PTP component protein, and block the translocation of cytochrome c from mitochondria to cytosol. On the other hand, pre-incubation with 25, 50 and 100 microg mL(-1) AA inhibited the Ca(2+)-induced mitochondrial permeability transition (MPT), including mitochondrial swelling, membrane potential dissipation and releasing of matrix Ca(2+) in liver mitochondria separated from normal mice, indicating the direct role of AA on mitochondria. Collectively, the above data suggest that AA could protect liver from damage and the mechanism might be related to up-regulating mitochondrial VDACs and inhibiting the process of MPT.

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Calcium; Chemical and Drug Induced Liver Injury; Cytochromes c; Galactosamine; Ion Channels; Lipopolysaccharides; Liver; Male; Membrane Potentials; Mice; Mice, Inbred ICR; Mitochondria, Liver; Mitochondrial Membrane Transport Proteins; Mitochondrial Permeability Transition Pore; Mitochondrial Swelling; Pentacyclic Triterpenes; RNA, Messenger; Triterpenes; Voltage-Dependent Anion Channels