• Asian-Australasian Journal of Animal Sciences
• /
• v.24 no.6
• /
• pp.857-866
• /
• 2011

This study examined the metabolism of free radicals in hepatic mitochondria of goats induced by lipopolysaccharide (LPS), and investigated the effects of melatonin (MT). Forty-eight healthy goats ($10{\pm}1.2\;kg$) were randomly selected and divided into four groups: saline control, LPS, MT+LPS and MT. The goats within each group were3 sacrificed either 3 or 6 h after treatment and the livers removed to isolate mitochondria. The respiration control ratio (RCR), the ADP:O ratio, the oxidative phosphorylation ratio (OPR), the concentration of $H_2O_2$ and the activities of Complex I-IV were determined. The mitochondrial membrane potential ($\Delta\psi_m$) was analyzed by flow cytometry. The results showed that RCR, O/P and OPR of the LPS group decreased (p<0.05), as well as activities of respiratory complexes, whereas the generation of $H_2O_2$ in Complex III increased (p<0.05) after 3 h, while Complex II and III increased after 6 h. Also, it was found that the mitochondrial membrane potential of the LPS group declined (p<0.05). However, pre-treatment with MT attenuated the injury induced by LPS, which not only presented higher (p<0.05) RCR, O/P, OPR, and respiratory complex activities, but also maintained the $\Delta\psi_m$. Interestingly, it is revealed that, in the MT+LPS group, the generation of $H_2O_2$ increased firstly in 3 h, and then significantly (p<0.05).decreased after 6 h. In the MT group, the function of mitochondria, the transmenbrane potential and the generation of $H_2O_2$ were obviously improved compared to the control group. Conclusion: melatonin prevents damage caused by LPS on hepatic mitochondria of goats.

• The Korean Journal of Physiology and Pharmacology
• /
• v.22 no.3
• /
• pp.311-319
• /
• 2018

Mitochondrial calcium overload is a crucial event in determining the fate of neuronal cell survival and death, implicated in pathogenesis of neurodegenerative diseases. One of the driving forces of calcium influx into mitochondria is mitochondria membrane potential (${\Delta}{\psi}_m$). Therefore, pharmacological manipulation of ${\Delta}{\psi}_m$ can be a promising strategy to prevent neuronal cell death against brain insults. Based on these issues, we investigated here whether nobiletin, a Citrus polymethoxylated flavone, prevents neurotoxic neuronal calcium overload and cell death via regulating basal ${\Delta}{\psi}_m$ against neuronal insult in primary cortical neurons and pure brain mitochondria isolated from rat cortices. Results demonstrated that nobiletin treatment significantly increased cell viability against glutamate toxicity ($100{\mu}M$, 20 min) in primary cortical neurons. Real-time imaging-based fluorometry data reveal that nobiletin evokes partial mitochondrial depolarization in these neurons. Nobiletin markedly attenuated mitochondrial calcium overload and reactive oxygen species (ROS) generation in glutamate ($100{\mu}M$)-stimulated cortical neurons and isolated pure mitochondria exposed to high concentration of $Ca^{2+}$ ($5{\mu}M$). Nobiletin-induced partial mitochondrial depolarization in intact neurons was confirmed in isolated brain mitochondria using a fluorescence microplate reader. Nobiletin effects on basal ${\Delta}{\psi}_m$ were completely abolished in $K^+-free$ medium on pure isolated mitochondria. Taken together, results demonstrate that $K^+$ influx into mitochondria is critically involved in partial mitochondrial depolarization-related neuroprotective effect of nobiletin. Nobiletin-induced mitochondrial $K^+$ influx is probably mediated, at least in part, by activation of mitochondrial $K^+$ channels. However, further detailed studies should be conducted to determine exact molecular targets of nobiletin in mitochondria.

• Journal of Microbiology
• /
• v.41 no.1
• /
• pp.41-45
• /
• 2003

N crassa mutant strain nap showed reduced growth rate, decreased electric membrane potential, and elevated intracellular ATP content in comparison to the wild type. Blue light induced a hyperpolarization of the membrane potential in both strains. The analysis of oxidative and phosphorylation activities of mitochondria isolated from the two strains has revealed that nap utilized more efficient oxidative pathways. The higher intracellular ATP content in the nap was presumably due to impaired transport systems of the plasma membrane, and to a lesser extent to the functioning of the fully competent respiratory chain. The excess ATP possibly accounts for carotenoid accumulation in the mutant.

• Proceedings of the PSK Conference
• /
• 2003.10b
• /
• pp.167.1-167.1
• /
• 2003

Here we show that panaxadiol, a ginseng saponin with a dammarane skeleton, induces acute apoptotic cell death in human hepatoma SK-HEP-1 cells as evidenced by analysis of DNA fragmentation, caspase activation, and changes in cell morphology. The kinetic study showed that panaxadiol-induced apoptosis is associated with depolarization of mitochondrial membrane potential and cytochrome c release. Sequential activations of caspases-depolarization of mitochondrial membrane potential and cytochrome c release. Sequential activations of caspases-9, and -3, or -7, but not of caspase 8 coincide well in a time dependent manner with mitochondrial membrane depolarization and cytochrome c release from mitochondria during apoptosis of SK-HEP-1 cells induced by treatment with panaxadiol. (omitted)

• Applied Biological Chemistry
• /
• v.28 no.4
• /
• pp.271-277
• /
• 1985

A polarograph with specially designed cell compartment usable in kinetic study of the mitochondrial respiration of a small sized sample was made, and its performance and experimental conditions were examined. An applied potential (ca-1.2V vs. SCE) which gives rise to the second step reduction of oxygen caused a considerable level of a residual current independent of oxygen, which is temporarily interpreted as the reduction current of the membrane-bound redox material(s) of mitochondria. A potential corresponding to the first slop reduction of oxygen (ca-0.4V vs SCE) did not produce the residual current. Thus, it is suggested that a measurement of oxygen concentration in a sample of mitochondria and submitochondrial particles by using dropping mercury electrode should be done with an applied potential of about -0.4V vs SCE. Consumption of oxygen by mitochondria was observed to follow practically zero order kinetics. Its rate constant exhibited the proportional relationship with the respiratory activity of mitochondria. Usefulness of tile instrument was properly demonstrated in the work on the temperature effect on the respiration of mitochondria isolated from several plant 4issues which were selected on the basis of chilling susceptivity.

• BMB Reports
• /
• v.42 no.11
• /
• pp.719-724
• /
• 2009

Recent studies have revealed that endoplasmic reticulum (ER) disturbance is involved in the pathophysiology of neurodegenerative disorders, contributing to the activation of the ER stress-mediated apoptotic pathway. Therefore, we investigated here the molecular mechanisms underlying the ER-mitochondria axis, focusing on calcium as a potential mediator of cell death signals. Using NT2 cells treated with brefeldin A or tunicamycin, we observed that ER stress induces changes in the mitochondrial function, impairing mitochondrial membrane potential and distressing mitochondrial respiratory chain complex Moreover, stress stimuli at ER level evoked calcium fluxes between ER and mitochondria. Under these conditions, ER stress activated the unfolded protein response by an overexpression of GRP78, and also caspase-4 and-2, both involved upstream of caspase-9. Our findings show that ER and mitochondria interconnection plays a prominent role in the induction of neuronal cell death under particular stress circumstances.

• The Journal of Korean Obstetrics and Gynecology
• /
• v.18 no.1
• /
• pp.29-44
• /
• 2005

To address the ability of Herba Portulacae(HP) to induce cell death, we investigated the effect of HP on cell viability. Twenty-four hours later, loss of viability occurred following HP exposure in a dose-dependent manner. The treatment of HP, a commonly used herb formulation in Korea, Japan and China, caused a decrease in cell viability. HP also resulted in apoptotic morphology a brightly blue-fluorescent condensed nuclei by Hoechst 33258-staining, and reduction of cell volume. Our results show that 2mg/ml HP induces mitochondria membrane potential collapse. Immunoblotting data also shows that the expression of Bcl-2, antiaoptotic protein, decrease by the addition of HP. This GFP-Bax overexpression system shows that an important pro-apoptotic Bcl-2-family protein, Bax is translocated to mitochondria by the addition of 2mg/ml HP. Inerestingly, MAPK inhibitor study shows that p38 MAPK inhibitor, SB203580 inhibits HP-induced cell death and caspase-3 activation in HP-treated HeLa cells. Furthermore, HP transiently but significantly induces p38 activation. But P38 MAPK inhibitor does not have any effect on the translocation of Bax. Considering these results, HP induces apoptosis via p38 MAPK activation. But the pathway does not involve the translocation of Bax.

• Biomolecules & Therapeutics
• /
• v.22 no.6
• /
• pp.519-524
• /
• 2014

We have previously shown that 2,4,3',5'-tetramethoxystilbene (TMS), a trans-stilbene analogue, induces apoptosis in human cancer cells. However, the detailed mechanisms of mitochondria-dependent apoptosis induced by TMS are not fully understood. In the present study, the possible roles of annexin A5 in TMS-mediated apoptosis were investigated in MCF7 human breast cancer cells. Quantitative real-time PCR analysis and Western blot analysis showed that the expression of annexin A5 was strongly increased in TMS-treated cells. TMS caused a strong translocation of annexin A5 from cytosol into mitochondria. Confocal laser scanning microscopic analysis clearly showed that TMS induced translocation of annexin A5 into mitochondria. TMS increased the expression and oligomerization of voltage-dependent anion channel (VDAC) 1, which may promote mitochondria-dependent apoptosis through disruption of mitochondrial membrane potential. When cells were treated with TMS, the levels of Bax, and Bak as well as annexin A5 were strongly enhanced. Moreover, we found that the cytosolic release of apoptogenic factors such as cytochrome c, or apoptosis-inducing factor (AIF) in mitochondria was markedly increased. Annexin A5 depletion by siRNA led to decreased proapoptotic factors such as Bax, Bak, and annexin A5. Taken together, our results indicate that annexin A5 may play an important role in TMS-mediated mitochondrial apoptosis through the regulation of proapoptotic proteins and VDAC1 expression.

• Molecules and Cells
• /
• v.39 no.9
• /
• pp.680-686
• /
• 2016

Uncoupling proteins (UCPs) are mitochondrial inner membrane proteins that function to dissipate proton motive force and mitochondrial membrane potential. One UCP has been identified in Caenorhabditis elegans (C. elegans), namely UCP-4. In this study, we examined its expression and localization using a GFP marker in C. elegans. ucp-4 was expressed throughout the body from early embryo to aged adult and UCP-4 was localized in the mitochondria. It is known that increased mitochondrial membrane protential leads to a reactive oxygen species (ROS) increase, which is associated with age-related diseases, including neurodegenerative diseases in humans. A ucp-4 mutant showed increased mitochondrial membrane protential in association with increased neuronal defects during aging, and the neurons of ucp-4 overexpressing animals showed decreased neuronal defects during aging. These results suggest that UCP-4 may be involved in neuroprotection during aging via relieving mitochondrial membrane protential. We also investigated the relationship between UCP-4 and innate immunity because increased ROS can affect innate immunity. ucp-4 mutant displayed increased resistance to the pathogen Staphylococcus aureus compared to wild type. The enhanced immunity in the ucp-4 mutant could be related to increased mitochondrial membrane protential, presumably followed by increased ROS. In summary, UCP-4 might have an important role in neuronal aging and innate immune responses through mediating mitochondrial membrane protential.

• Journal of Microbiology and Biotechnology
• /
• v.23 no.11
• /
• pp.1627-1635
• /
• 2013

Mitochondria often play central roles in apoptotic pathways, and disruption of the mitochondrial transmembrane potential (${\Delta}{\psi}m$) has been observed in various cells undergoing apoptosis. Human cytomegalovirus (HCMV) infection induces apoptosis in permissive cells; however, investigations of mitochondria-targeted apoptosis in HCMV-infected human foreskin fibroblast (HFF) cells have been limited. Here, we investigated the mitochondrial apoptosis pathway in HCMV-infected HFF cells. Flow cytometry analysis using JC-1 revealed that HCMV infection induces disruption of ${\Delta}{\psi}m$ in HFF cells when administered 24 h post-infection (hpi), and this disruption was maximized at 48 hpi. Moreover, cytochrome c, normally a mitochondrial inner membrane protein, was detected in cytoplasmic extracts of HCMV-infected cells, but not mock-infected cells, by western blot analysis at 24 hpi. A caspase activity assay based on fluorescence spectrophotometry using a fluorogenic substrate revealed an increase in caspase-3 activity at 48 hpi in HCMV-infected cells. Caspase-8 activity was increased at 72 hpi in HCMV-infected cells. These results imply that HCMV infection induces mitochondria-mediated apoptosis in HFF cells.