• 한국생물공학회:학술대회논문집
• /
• 2005.10a
• /
• pp.205-205
• /
• 2005

• The Korean Journal of Physiology and Pharmacology
• /
• v.10 no.1
• /
• pp.51-58
• /
• 2006

The promoting effect of hydrogen peroxide ($H_2O_2$) against the cytotoxicity of 1-methyl-4-phenylpyridinium ($MPP^+$) in differentiated PC12 cells was assessed by measuring the effect on the mitochondrial membrane permeability. Treatment of PC12 cells with $MPP^+$ resulted in the nuclear damage, decrease in the mitochondrial transmembrane potential, cytosolic accumulation of cytochrome c, activation of caspase-3, increase in the formation of reactive oxygen species (ROS) and depletion of GSH. Addition of $H_2O_2$ enhanced the $MPP^+-induced$ nuclear damage and cell death. Catalase, Carboxy-PTIO, Mn-TBAP, N-acetylcysteine, cyclosporin A and trifluoperazine inhibited the cytotoxic effect of $MPP^+$ in the presence of $H_2O_2$. Addition of $H_2O_2$ promoted the change in the mitochondrial membrane permeability, ROS formation and decrease in GSH contents due to $MPP^+$ in PC12 cells. The results show that the $H_2O_2$ treatment promotes the cytotoxicity of $MPP^+$ against PC12 cells. $H_2O_2$ may enhance the $MPP^+$-induced viability loss in PC12 cells by promoting the mitochondrial membrane permeability change, release of cytochrome c and subsequent activation of caspase-3, which is associated with the increased formation of ROS and depletion of GSH. The findings suggest that $H_2O_2$ as a promoting agent for the formation of mitochondrial permeability transition may enhance the neuronal cell injury caused by neurotoxins.

• Journal of Ginseng Research
• /
• v.47 no.2
• /
• pp.199-209
• /
• 2023

Background: Due to the interrupted blood supply in cerebral ischemic stroke (CIS), ischemic and hypoxia results in neuronal depolarization, insufficient NAD+, excessive levels of ROS, mitochondrial damages, and energy metabolism disorders, which triggers the ischemic cascades. Currently, improvement of mitochondrial functions and energy metabolism is as a vital therapeutic target and clinical strategy. Hence, it is greatly crucial to look for neuroprotective natural agents with mitochondria protection actions and explore the mediated targets for treating CIS. In the previous study, notoginseng leaf triterpenes (PNGL) from Panax notoginseng stems and leaves was demonstrated to have neuroprotective effects against cerebral ischemia/reperfusion injury. However, the potential mechanisms have been not completely elaborate. Methods: The model of middle cerebral artery occlusion and reperfusion (MCAO/R) was adopted to verify the neuroprotective effects and potential pharmacology mechanisms of PNGL in vivo. Antioxidant markers were evaluated by kit detection. Mitochondrial function was evaluated by ATP content measurement, ATPase, NAD and NADH kits. And the transmission electron microscopy (TEM) and pathological staining (H&E and Nissl) were used to detect cerebral morphological changes and mitochondrial structural damages. Western blotting, ELISA and immunofluorescence assay were utilized to explore the mitochondrial protection effects and its related mechanisms in vivo. Results: In vivo, treatment with PNGL markedly reduced excessive oxidative stress, inhibited mitochondrial injury, alleviated energy metabolism dysfunction, decreased neuronal loss and apoptosis, and thus notedly raised neuronal survival under ischemia and hypoxia. Meanwhile, PNGL significantly increased the expression of nicotinamide phosphoribosyltransferase (NAMPT) in the ischemic regions, and regulated its related downstream SIRT1/2/3-MnSOD/PGC-1α pathways. Conclusion: The study finds that the mitochondrial protective effects of PNGL are associated with the NAMPT-SIRT1/2/3-MnSOD/PGC-1α signal pathways. PNGL, as a novel candidate drug, has great application prospects for preventing and treating ischemic stroke.

• The Korean Journal of Physiology and Pharmacology
• /
• v.11 no.2
• /
• pp.37-43
• /
• 2007

Adenosine has been reported to provide cytoprotection in the central nervous systems as well as myocardium by activating cell surface adenosine receptors. However, the exact target and mechanism of its action still remain controversial. The present study was performed to examine whether adenosine has a protective effect against $A{\beta}$-induced injury in neuroglial cells. The astrocyte-derived human neuroglioma cell line, A172 cells, and $A{\beta}_{25{\sim}35}$ were employed to produce an experimental $A{\beta}$-induced glial cell injury model. Adenosine significantly prevented $A{\beta}$-induced apoptotic cell death. Studies using various nucleotide receptor agonists and antagonists suggested that the protection was mediated by $A_1$ receptors. Adenosine attenuated $A{\beta}$-induced impairment in mitochondrial functional integrity as estimated by cellular ATP level and MTT reduction ability. In addition, adenosine prevented $A{\beta}$-induced mitochondrial permeability transition, release of cytochrome c into cytosol and subsequent activation of caspase-9. The protective effect of adenosine disappeared when cells were pretreated with 5-hydroxydecanoate, a selective blocker of the mitochondrial ATP-sensitive $K^+$ channel. In conclusion, therefore we suggest that adenosine exerts protective effect against $A{\beta}$-induced cell death of A172 cells, and that the underlying mechanism of the protection may be attributed to preservation of mitochonarial functional integrity through opening of the mitochondrial ATP-sensitive $K^+$ channels.

• The Korean Journal of Pharmacology
• /
• v.32 no.2
• /
• pp.201-209
• /
• 1996

Restoration of the blood flow after a period of ischemia is accompanied by generation of toxic oxygen radicals. This phenomenon may account for the occurrence of reperfusion-mediated tissue injury in ischemic hearts. In in vitro studies, although oxygen radicals can be generated from a variety of sources, including xanthine oxidase system, activated leucocytes, mitochondria and others, the most important source and mechanism of oxygen radical production in the post-ischemic reperfused hearts is unclear. In the present study, we tested the hypothesis that the respiratory chain of mitochondria might be an important source of oxygen radicals which are responsible for the development of the reperfusion injury of ischemic hearts. Langendorff-perfused, isolated rat hearts were subjected to 30 min of global ischemia at $37^{\circ}C$, followed by reperfusion. Amytal, a reversible inhibitor of mitochondrial respiration, was employed to assess the mitochondrial contributions to the development of the reperfusion injury. Intact mitochonria were isolated from the control and the post-ischemic reperfused hearts. Mitochondrial oxygen radical generation was measured by chemiluminescence method and the oxidative tissue damage was estimated by measuring a lipid peroxidation product, malondialdehyde(MDA). To evaluate the extent of the reperfusion injury, post-ischemic functional recovery and lactate dehydrogenase(LDH) release were assessed and compared in Amytal-treated and -untreated hearts. Upon reperfusion of the ischemic hearts, MDA release into the coronary effluent was markedly increased. MDA content of mitochondria isolated from the post-ischemic reperfused hearts was increased to 152% of preischemic value, whereas minimal change was observed in extramitochondrial fraction. The generation of superoxide anion was increased about twice in mitochondria from the reperfused hearts than in those from the control hearts. Amytal inhibited the mitochondrial superoxide generation significantly and also suppressed MDA production in the reperfused hearts. Additionally, Amytal prevented the contractile dysfunction and the increased release of LDH observed in the reperfused hearts. In conclusion, these results indicate that the respiratory chain of mitochondria may be an important source of oxygen radical formation in post-ischemic reperfused hearts, and that oxygen radicals originating from the mitochondria may contribute to the development of myocardial reperfusion injury.

• Biomolecules & Therapeutics
• /
• v.31 no.1
• /
• pp.97-107
• /
• 2023

Aristolochic acid (AA), extracted from Aristolochiaceae plants, plays an essential role in traditional herbal medicines and is used for different diseases. However, AA has been found to be nephrotoxic and is known to cause aristolochic acid nephropathy (AAN). AA-induced acute kidney injury (AKI) is a syndrome in AAN with a high morbidity that manifests mitochondrial damage as a key part of its pathological progression. Melatonin primarily serves as a mitochondria-targeted antioxidant. However, its mitochondrial protective role in AA-induced AKI is barely reported. In this study, mice were administrated 2.5 mg/kg AA to induce AKI. Melatonin reduced the increase in Upro and Scr and attenuated the necrosis and atrophy of renal proximal tubules in mice exposed to AA. Melatonin suppressed ROS generation, MDA levels and iNOS expression and increased SOD activities in vivo and in vitro. Intriguingly, the in vivo study revealed that melatonin decreased mitochondrial fragmentation in renal proximal tubular cells and increased ATP levels in kidney tissues in response to AA. In vitro, melatonin restored the mitochondrial membrane potential (MMP) in NRK-52E and HK-2 cells and led to an elevation in ATP levels. Confocal immunofluorescence data showed that puncta containing Mito-tracker and GFP-LC3A/B were reduced, thereby impeding the mitophagy of tubular epithelial cells. Furthermore, melatonin decreased LC3A/B-II expression and increased p62 expression. The apoptosis of tubular epithelial cells induced by AA was decreased. Therefore, our findings revealed that melatonin could prevent AA-induced AKI by attenuating mitochondrial damage, which may provide a potential therapeutic method for renal AA toxicity.

• Journal of Ginseng Research
• /
• v.47 no.3
• /
• pp.408-419
• /
• 2023

Background: Ginsenoside compound K (CK), the main active metabolite in Panax ginseng, has shown good safety and bioavailability in clinical trials and exerts neuroprotective effects in cerebral ischemic stroke. However, its potential role in the prevention of cerebral ischemia/reperfusion (I/R) injury remains unclear. Our study aimed to investigate the molecular mechanism of ginsenoside CK against cerebral I/R injury. Methods: We used a combination of in vitro and in vivo models, including oxygen and glucose deprivation/reperfusion induced PC12 cell model and middle cerebral artery occlusion/reperfusion induced rat model, to mimic I/R injury. Intracellular oxygen consumption and extracellular acidification rate were analyzed by Seahorse multifunctional energy metabolism system; ATP production was detected by luciferase method. The number and size of mitochondria were analyzed by transmission electron microscopy and MitoTracker probe combined with confocal laser microscopy. The potential mechanisms of ginsenoside CK on mitochondrial dynamics and bioenergy were evaluated by RNA interference, pharmacological antagonism combined with co-immunoprecipitation analysis and phenotypic analysis. Results: Ginsenoside CK pretreatment could attenuate mitochondrial translocation of DRP1, mitophagy, mitochondrial apoptosis, and neuronal bioenergy imbalance against cerebral I/R injury in both in vitro and in vivo models. Our data also confirmed that ginsenoside CK administration could reduce the binding affinity of Mul1 and Mfn2 to inhibit the ubiquitination and degradation of Mfn2, thereby elevating the protein level of Mfn2 in cerebral I/R injury. Conclusion: These data provide evidence that ginsenoside CK may be a promising therapeutic agent against cerebral I/R injury via Mul1/Mfn2 mediated mitochondrial dynamics and bioenergy.

• Proceedings of the Plant Resources Society of Korea Conference
• /
• 2021.04a
• /
• pp.57-57
• /
• 2021

Oxygen glucose deprivation/re-oxygenation (OGD/R) induces neuronal injury via mechanisms that are believed to mimic the pathways associated with brain ischemia. Stachys sieboldii Miq. (Chinese artichoke), which has been extensively used in oriental traditional medicine to treat of ischemic stroke; however, the role of S. sieboldii Miq. (SSM) in OGD/R induced neuronal injury is not yet fully understood. The present research is aimed to investigate the protective effect and possible mechanisms of SSM extract treatment in an in vitro model of OGD/R to simulate ischemia/reperfusion Injury. Pretreatment of these cells with SSM significantly attenuated OGD/R-induced production of reactive oxygen species (ROS) by increasing GPx, SOD, and decreasing MDA. SSM decreased mitochondrial damage caused by OGD/R injury and inhibited the release of cyt-c from mitochondrion to cytoplasm in SH-SY5Y cells. Furthermore, neuronal cell apoptosis caused by OGD/R injury was inhibited by SSM, and SSM could decrease apoptosis by increasing ratio of Bcl-2/Bax and inhibiting caspase signaling pathway in SH-SY5Y cells. SSM demonstrated a neuroprotective effect on the simulated cerebral ischemia in vitro model, and this effect was the inhibition of mitochondria-mediated apoptosis pathway by scavenging of ROS generation. Therefore, SSM may be a promising neuroprotective strategy against ischemic stroke.

• Korean Journal of Veterinary Research
• /
• v.64 no.1
• /
• pp.2.1-2.8
• /
• 2024

This study was performed to assess the antiapoptotic effect of canine platelet-rich plasma (PRP) treated on the canine adipose-derived mesenchymal stem cells (cMSCs) under cold ischemic conditions. The effect of preventing apoptosis of cMSCs was evaluated in the apoptotic condition induced by cold ischemic injury in vitro. To determine the progression of apoptosis, the changes in cell nucleus were observed using 4',6-diamidino-2-phenylindole (DAPI) fluorescence staining. In addition, we examined the mitochondrial membrane potential (MMP) and caspase-3 activity. When the cold hypoxic injury was applied to cMSCs, the apoptotic change was observed by DAPI staining, mitochondrial staining for MMP, and caspase-3 assay. PRP significantly decreased the number of apoptotic cells. Nuclear shrinkage and fragmentation of apoptotic cells in control groups were observed by DAPI staining. The MMP was recovered by the treatment of PRP. In addition, when the luminescence intensity was measured for caspase-3 activity, the value was significantly higher in the PRP treated groups than the control groups. The results of this study showed that the PRP may have a beneficial effect on apoptosis induced by cold ischemic injury.

• Journal of Nutrition and Health
• /
• v.48 no.5
• /
• pp.390-397
• /
• 2015

Purpose: In this study, we investigated the effects of chronic alcohol and excessive iron intake on mitochondrial DNA (mtDNA) damage and the progression of alcoholic liver injury in rats. Methods: Twenty-four Sprague-Dawley male rats were divided into four groups (Control, EtOH, Fe, and EtOH + Fe), and fed either control or ethanol (36% of total calories) liquid diet with or without 0.6% carbonyl iron for eight weeks. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, liver malondialdehyde concentrations were measured by colorimetric assays. Liver histopathology was examined by Hematoxylin-eosin staining of the fixed liver tissues. The integrity of the hepatic mtDNA and nuclear DNA was measured by long-range PCR. The gene expression levels of cytochrome c oxidase subunit 1 (Cox1) and NADH dehydrogenase subunit 4 (Nd4) were examined by real-time PCR. Results: Serum ALT and AST activities were significantly higher in the EtOH+Fe group, as compared to the Control group. Similarly, among four groups, liver histology showed the most severe lipid accumulation, inflammation, and necrosis in the EtOH + Fe group. PCR amplification of near-full-length (15.9 kb) mtDNA showed more than 50% loss of full-length product in the liver of the EtOH + Fe group, whereas amounts of PCR products of a nuclear DNA were unaffected. In addition, the changes in the mtDNA integrity showed correlation with reductions in the mRNA levels of mitochondrial gene Cox1 and Nd4. Conclusion: Our data suggested that the liver injury associated with excessive iron and alcohol intake involved mtDNA damage and corresponding mitochondrial dysfunction.