• Clinical and Experimental Reproductive Medicine
• /
• 제48권4호
• /
• pp.316-321
• /
• 2021

Objective: Amino acids can protect sperm structure in cryopreservation due to their antioxidant properties. Therefore, the present study aimed to investigate the protective effect of L-carnitine (LC) and N-acetyl cysteine (NAC) on motility parameters, plasma membrane integrity (PMI), mitochondrial membrane potential (MMP), DNA damage, and human sperm intracellular reactive oxygen species (ROS) during vitrification. Methods: Twenty normal human sperm samples were examined. Each sample was divided into six equal groups: LC (1 and 10 mM), NAC (5 and 10 mM), and cryopreserved and fresh control groups. Results: The groups treated with LC and NAC showed favorable findings in terms of motility parameters, DNA damage, and MMP. Significantly higher levels of intracellular ROS were observed in all cryopreserved groups than in the fresh group (p≤0.05). The presence of LC and NAC at both concentrations caused an increase in PMI, MMP, and progressive motility parameters, as well as a significant reduction in intracellular ROS compared to the control group (p≤0.05). The concentrations of the amino acids did not show any significant effect. Conclusion: LC and NAC are promising as potential additives in sperm cryopreservation.

• Toxicological Research
• /
• 제35권1호
• /
• pp.83-91
• /
• 2019

Nanoparticles (NPs) have been recognized as both useful tools and potentially toxic materials in various industrial and medicinal fields. Previously, we found that zinc oxide (ZnO) NPs that are neurotoxic to human dopaminergic neuroblastoma SH-SY5Y cells are mediated by lipoxygenase (LOX), not cyclooxygenase-2 (COX-2). Here, we examined whether human bone marrow-derived mesenchymal stem cells (MSCs), which are different from neuroblastoma cells, might exhibit COX-2- and/or LOX-dependent cytotoxicity of ZnO NPs. Additionally, changes in annexin V expression, caspase-3/7 activity, and mitochondrial membrane potential (MMP) induced by ZnO NPs and ZnO were compared at 12 hr and 24 hr after exposure using flow cytometry. Cytotoxicity was measured based on lactate dehydrogenase activity and confirmed by trypan blue staining. Rescue studies were executed using zinc or iron chelators. ZnO NPs and ZnO showed similar dose-dependent and significant cytotoxic effects at concentrations ${\geq}15{\mu}g/mL$, in accordance with annexin V expression, caspase-3/7 activity, and MMP results. Human MSCs exhibited both COX-2 and LOX-mediated cytotoxicity after exposure to ZnO NPs, which was different from human neuroblastoma cells. Zinc and iron chelators significantly attenuated ZnO NPs-induced toxicity. Conclusively, these results suggest that ZnO NPs exhibit both COX-2- and LOX-mediated apoptosis by the participation of mitochondrial dysfunction in human MSC cultures.

• International Journal of Oral Biology
• /
• 제34권4호
• /
• pp.185-190
• /
• 2009

This study examined the effects of red light generated from a light emitting diode (LED) upon proliferation and mitochondrial stress in human gingival fibroblasts (hGFs). Cells were exposed to LED-generated red light at a clinically relevant intensity and distance with a 610-630 nm wavelength for various times (0-48 min). At different exposure times, cells were processed for the analysis of succinate dehydrogenase (SDH) activity, proliferation, mitochondrial membrane potential (MMP) and cytotoxicity. Cell cycle progression was also investigated by flow cytometry after staining with propidium iodide. Red light exposure was found to inhibit SDH activity and DNA synthesis in hGFs in a time-dependent manner. Light exposure also reduced the MMP levels in these cells and this was closely associated with a $G_0/G_1$ arrest. In contrast, exposure of hGFs to red light for 48 min led to a dramatic loss of MMP with an attendant increase in cytotoxicity. These findings demonstrate that LED-generated red light may cause mitochondrial stress and growth inhibition in hGFs during tooth whitening therapy, depending on the length of the exposure.

• 대한수의학회지
• /
• 제64권1호
• /
• 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.

• 생명과학회지
• /
• 제19권5호
• /
• pp.598-606
• /
• 2009

본 연구는 저산소증에서 반하가 대뇌신경세포에 미치는 영향을 알아보기 위하여 $E_18$의 배양 흰쥐 대뇌신경세포를 반하로 전처리한 후, LDH assay와 tryphan blue 염색으로 세포 생존율을 측정하였고, $H_2DCF-DA$, JC-1 염색으로 MMP, ROS 및 RNS 변화를 조사하였다. 이에 반하는 저산소증으로 유발된 대뇌신경세포를 2.5 ${\mu}g/ml$까지 농도의존적으로 세포 생존률을 증가시켰으며, 시간에 따른 생존율을 살펴보면 저산소증 유발 후 1 시간에는 별 차이를 보이지 않았지만 3 일, 5 일 후에는 각각 10.2%, 17.8%로 매우 유의한 증가를 보였다. 저산소증에서 반하가 MMP에 미치는 영향을 보기 위해 저산소증 유발직전과 유발 후 1 일, 3 일, 5 일에 JC-1으로 염색하고 미토콘드리아의 염색강도를 측정한 결과 적색형광은 실험군에서 전반적으로 대조군에 비하여 강하게 염색되는 미토콘드리아의 비율을 증가시킨 반면 녹색형광은 대조군과 뚜렷한 차이를 보이지 않았다. 즉 반하가 저산소증으로 유발된 MMP의 소실을 감소시킴을 알 수 있다. 또한 반하는 전반적으로 $H_2DCF-DA$에 염색되는 세포 비율을 현저하게 낮추는 것으로 나타나 저산소증으로 유발된 ROS 및 RNS의 생성을 유의성 있게 감소시켰다. 따라서 반하는 저산소증에서 ROS의 생성을 억제하고 MMP의 소실을 막아 세포의 에너지고갈을 방지함으로서 신경세포를 보호하는 것으로 이해된다.

• 대한본초학회지
• /
• 제36권1호
• /
• pp.41-49
• /
• 2021

Objective : Citrus unshiu peel (Citri Unshius Pericarpium) has been prescribed to suppress coughing and phlegm in Korean medicine. In this study, the effect of ethanol extract of Citrus unshiu peel (CEE) on apoptosis was investigated using cadmium chloride (CdCl2) treated HepG2 cells. Methods : CEE was prepared by extracting 300 g of Citri Unshius Pericarpium in 3 L of ethanol for 72 h. Apoptosis was determined by the TUNEL assay. The mitochondrial membrane potential (MMP) was monitored using the membrane-permeable fluorescent dye Rh123. The expression level of each protein was monitored by Western blot analysis. Results : CEE protected HepG2 cells from apoptosis as determined by the TUNEL assay. A decrease in MMP was observed in cells exposed to cadmium, indicating that mitochondria are involved in the induction of apoptosis. However, CEE recovered the reduction in MMP caused by cadmium. In addition, decreased expression of B-cell lymphoma 2 (Bcl-2), procaspase, and poly(ADP-ribose) polymerase (PARP) by cadmium was increased by CEE. The anti-apoptotic effect of CEE was found to be associated with inhibition of JNK and p38 phosphorylation when examining the expression of phosphorylated MAPK by Western blot. Conclusion : This study showed that CEE exerted anti-apoptotic effects in cadmium-induced HepG2 cells by inhibiting the reduction of MMP and changes in the expression level of apoptotic proteins. These results suggest the potential for CEE to be used for heavy metal-induced liver damage.

• Clinical and Experimental Reproductive Medicine
• /
• 제49권2호
• /
• pp.110-116
• /
• 2022

Objective: Sperm vitrification leads to the production of reactive oxygen species (ROS) that can damage the functional parameters of sperm. The present study aimed to investigate the antioxidant effect of Nigella sativa extract on motility, plasma membrane function, mitochondrial membrane potential (MMP), DNA damage, and intracellular ROS production. Methods: A total of 20 sperm samples were used. Samples were divided into six experimental groups, including groups with aqueous extract from N. sativa seeds at concentrations of 1% to 6%, a cryopreserved control group, and a fresh control group. Results: Statistical analysis showed significantly higher total sperm motility at concentrations of 3% to 6% than in the vitrified semen control group. Additionally, progressive motility and all motion characteristics at all concentrations were significantly higher than in the vitrified semen control group. The presence of N. sativa seed extract also improved the quality of the sperm parameters assayed in all experimental groups (1%-6%; intracellular ROS production, DNA damage, MMP, and sperm membrane function) compared to the control group. Conclusion: Higher concentrations of N. sativa led to improvements in all sperm parameters and sperm quality. These findings indicate that N. sativa seed extract is effective for improving the quality of sperm after vitrification.

• BMB Reports
• /
• 제43권1호
• /
• pp.57-61
• /
• 2010

Activator protein-1 can induce either cell survival or death, which is controlled by opposing effects of different Jun members. It is generally accepted that c-Jun is pro-apoptotic, but that JunD is anti-apoptotic in stress-exposed cells. Additionally, although there are reports suggesting that JunB plays a protective role, its role in stress-induced apoptosis remains unclear. Here, we investigated the role of JunB in $H_2O_2$-induced cell death using cells that over-expressed the protein or were transfected with si-JunB. Inhibition of JunB expression accelerated $H_2O_2$-mediated loss of mitochondrial membrane potential (MMP) and cytotoxicity. Conversely, over-expression of JunB protein led to significant inhibition of the MMP loss and cell death. The increase in JunB expression also attenuated nuclear relocation of apoptosis-inducing factor and mitochondrial Bcl-2 reduction that occurred following $H_2O_2$ exposure. These results suggest that JunB can signal survival against oxidant-mediated cell death by suppressing mitochondrial stress.

• Journal of Microbiology and Biotechnology
• /
• 제31권2호
• /
• pp.217-225
• /
• 2021

This study aimed to investigate the neuroprotective effects of 1-methoxylespeflorin G11 (MLG), a pterocarpan, against glutamate-induced neurotoxicity in neuronal HT22 hippocampal cells. The protective effects of MLG were evaluated using MTT assay and microscopic analysis. The extent of apoptosis was studied using flow cytometric analysis performed on the damaged cells probed with annexin V/propidium iodide. Moreover, mitochondrial reactive oxygen species (ROS) were assessed using flow cytometry through MitoSOXTM Red staining. To determine mitochondrial membrane potential, staining with tetramethylrhodamine and JC-1 was performed followed by flow cytometry. The results demonstrated that MLG attenuates glutamate-induced apoptosis in HT22 cells by inhibiting intracellular ROS generation and mitochondrial dysfunction. Additionally, MLG prevented glutamate-induced apoptotic pathway in HT22 cells through upregulation of Bcl-2 and downregulation of cleaved PARP-1, AIF, and phosphorylated MAPK cascades. In addition, MLG treatment induced HO-1 expression in HT22 cells. These results suggested that MLG exhibits neuroprotective effects against glutamate-induced neurotoxicity in neuronal HT22 cells by inhibiting oxidative stress and apoptosis.

• Journal of Microbiology and Biotechnology
• /
• 제18권11호
• /
• pp.1862-1867
• /
• 2008

Streptochlorin is a small molecule isolated from marine Streptomyces sp. that is known to have antiangiogenic and anticancer properties. In this study, we examined the effects of this compound on reactive oxygen species (ROS) production and the association of these effects with apoptotic tumor cell death, using a human hepatocarcinoma Hep3B cell line. The results of this study demonstrated that streptochlorin mediates ROS production, and that this mediation is followed by a decrease in the mitochondrial membrane potential (MMP, ${\Delta}{\Psi}_m$), activation of caspase-3, and downregulation of antiapoptotic Bcl-2 protein. The quenching of ROS generation by N-acetyl-L-cysteine administration, a scavenger of ROS, reversed the streptochlorin-induced apoptosis effects via inhibition of ROS production, MMP collapse, and the subsequent activation of caspase-3. These observations clearly indicate that ROS are involved in the early molecular events in the streptochlorin-induced apoptotic pathway. Taken together, our data imply that streptochlorin-induced ROS is a key mediator of MMP collapse, which leads to the caspase-3 activation, culminating in apoptosis.