• Yonsei Medical Journal
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• 제59권9호
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• pp.1096-1106
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• 2018

Purpose: Alzheimer's disease (AD) is the sixth most common cause of death in the United States. MicroRNAs have been identified as vital players in neurodegenerative diseases, including AD. microRNA-128 (miR-128) has been shown to be dysregulated in AD. This study aimed to explore the roles and molecular mechanisms of miR-128 in AD progression. Materials and Methods: Expression patterns of miR-128 and peroxisome proliferator-activated receptor gamma ($PPAR-{\gamma}$) messenger RNA in clinical samples and cells were measured using RT-qPCR assay. $PPAR-{\gamma}$ protein levels were determined by Western blot assay. Cell viability was determined by MTT assay. Cell apoptotic rate was detected by flow cytometry via double-staining of Annexin V-FITC/PI. Caspase 3 and $NF-{\kappa}B$ activity was determined by a Caspase 3 Activity Assay Kit or $NF-{\kappa}B$ p65 Transcription Factor Assay Kit, respectively. Bioinformatics prediction and luciferase reporter assay were used to investigate interactions between miR-128 and $PPAR-{\gamma}$ 3'UTR. Results: MiR-128 expression was upregulated and $PPAR-{\gamma}$ expression was downregulated in plasma from AD patients and $amyloid-{\beta}$ $(A{\beta})-treated$ primary mouse cortical neurons (MCN) and Neuro2a (N2a) cells. Inhibition of miR-128 decreased $A{\beta}-mediated$ cytotoxicity through inactivation of $NF-{\kappa}B$ in MCN and N2a cells. Moreover, $PPAR-{\gamma}$ was a target of miR-128. $PPAR-{\gamma}$ upregulation attenuated $A{\beta}-mediated$ cytotoxicity by inactivating $NF-{\kappa}B$ in MCN and N2a cells. Furthermore, $PPAR-{\gamma}$ downregulation was able to abolish the effect of anti-miR-128 on cytotoxicity and $NF-{\kappa}B$ activity in MCN and N2a cells. Conclusion: MiR-128 inhibitor decreased $A{\beta}-mediated$ cytotoxicity by upregulating $PPAR-{\gamma}$ via inactivation of $NF-{\kappa}B$ in MCN and N2a cells, providing a new potential target in AD treatment.

• Biomolecules & Therapeutics
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• 제17권1호
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• pp.70-78
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• 2009

Coptis japonica (C. japonica) is a perennial medicinal plant that has anti-inflammatory activity. C. japonica contains numerous biologically active alkaloids including berberine, palmatine, epi-berberine, and coptisine. The most well-known anti-inflammatory principal in C. japonica is berberine. For example, berberine has been implicated in the inhibition of iNOS induction by cytokines in microglial cells. However, the efficacies of other alkaloids components on microglial activation were not investigated yet. In this study, we investigated the effects of three alkaloids (palmatine, epi-berberine and coptisine) from C. japonica on lipopolysaccharide (LPS)-induced microglial activation. BV2 microglial cells were immunostimulated with LPS and then the production of several inflammatory mediators such as nitric oxide (NO), reactive oxygen species (ROS) and matrix metalloproteinase-9 (MMP-9) were examined as well as the phosphorylation status of Erk1/2 mitogen activated protein kinase (MAPK). Palmatine and to a lesser extent epi-berberine and coptisine, significantly reduced the release of NO, which was mediated by the inhibition of LPS-stimulated mRNA and protein induction of inducible nitric oxide synthase (iNOS) from BV2 microglia. In addition to NO, palmatine inhibited MMP-9 enzymatic activity and mRNA induction by LPS. Palmatine also inhibited the increase in the LPS-induced MMP-9 promoter activity determined by MMP-9 promoter luciferase reporter assay. LPS stimulation increased Erk1/2 phosphorylation in BV2 cells and these alkaloids inhibited the LPS-induced phosphorylation of Erk1/2. The anti-inflammatory effect of palmatine in LPS-stimulated microglia may suggest the potential use of the alkaloids in the modulation of neuroinflammatory responses, which might be important in the pathophysiological events of several neurological diseases including Alzheimer's disease (AD), multiple sclerosis (MS), Parkinson's disease (PD) and stroke.

• 생약학회지
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• 제45권1호
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• pp.55-61
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• 2014

Microwaves are non-ionizing electromagnetic waves of frequency between 300MHz to 300GHz and positioned between the X-ray and infrared rays in the electromagnetic spectrum. In recent years, the use of microwave for extraction of ingredient from plant material has shown remarkable research interest and potential. Scutellaria radix has been used as a traditional medicine for a variety of diseases. It has been reported to exert beneficial health effects, such as anti-bacterial, antiviral, anti-inflammatory, and free-radical scavenging. Oxidative stress or the accumulation of reactive oxygen species (ROS) leads neuronal cellular death and dysfunction, and it contributes to neuronal degenerative disease such as Alzheimer's disease, Parkinson's disease and stroke. In this study, we aimed to compare the neuroprotective and antioxidant effect of Scutellaria radix extracted by different methods using hot-water extraction (SBE-DW) or microwave extraction (SBE-DW-MW). As a result, we first examined HPLC analysis of hot-water and microwave extracts of Scutellaria radix. The hot-water and microwave extracts of Scutellaria radix showed the discernible difference patterns of HPLC analysis. Microwave-water extracts of Scutellaria radix increased DPPH radical scavenging activity more than hot-water extraction. Microwave-water extracts of Scutellaria radix also showed neuroprotective effects and ROS inhibition against glutamate-induced oxidative stress in mouse hippocampal HT22 cells, but hot-water extraction not showed. In addition, the phosphorylation of MAPKs induced by glutamate insult was prevented by microwave-water extracts of Scutellaria radix. Thus, these results suggested that microwave extraction can be utilized for improving the extraction efficiency and biological activity of Scutellaria radix.

• 동의생리병리학회지
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• 제22권5호
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• pp.1202-1209
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• 2008

Peroxynitrite ($ONOO^-$), superoxide anion radical (${\cdot}\;{O_2}^-$) and nitric oxide (NO) are cytotoxic because they can oxidize several cellular components such as proteins, lipids and DNA. They have been implicated in the aging processes, and age-related diseases such as Alzheimer's disease, rheumatoid arthritis, cancer, diabetes, obesity and atherosclerosis. The aim of this study was to investigate the effects of Ojunghwan on the generation of peroxynitrite ($ONOO^-$), nitric oxide (NO) and superoxide anion radical (${\cdot}\;{O_2}^-$), and on the expression of $NF-{\kappa}B$-dependent inflammatory proteins in ob/ob mice. Mice were grouped and treated for 5 weeks as follows. Both the normal lean (C57/BL6J black mice) and control obese (ob/ob mice) groups have received the standard chow. The experimental groups were fed with a diet of chow supplemented with 30 and 90 mg Ojung-hwan per 1 kg of body weight for 14 days. For this study, the fluorescent probes, namely 2',7'-dichlorodihydrofluorescein diacetate (DCFDA), 4,5-diaminofluorescein (DAF-2) and dihydrorhodamine 123 (DHR 123) were used. Western blot was performed using anti-phospho $I{\kappa}B-{\alpha}$, $anti-IKK-{\alpha}$, $anti-NF-{\kappa}B$ (p50, p65), anti-COX-2, anti-iNOS, anti-VCAM-1 and anti-MMP-9 antibodies, respectively. Ojunghwan inhibited the generation of $ONOO^-$, NO and ${\cdot}\;{O_2}^-$ in the lipopolysaccharide (LPS)-treated mouse kidney postmitochondrial fraction in vitro. The generation of $ONOO^-$, NO, ${\cdot}\;{O_2}^-$ and $PGE_2$ were inhibited in the Ojunghwan-administered ob/ob mice groups. The GSH/GSSG ratio was decreased in the ob/ob mice, whereas that were improved in the Ojunghwan-administered groups. Ojunghwan inhibited the expression of $phospho-I{\kappa}B-{\alpha}$, $IKK-{\alpha}$, $NF-{\kappa}B$ (p50, p65), COX-2, iNOS, VCAM-1 and MMP-9 genes. These results suggest that Ojunghwan is an effective scavenger of $ONOO^-$, ${\cdot}\;{O_2}^-$, NO and $PGE_2$, and has an inhibitory effect on the expression of $NF-{\kappa}B$-dependent inflammatory genes in ob/ob mice. Therefore, Ojunghwan might be used as a potential therapeutic drug against the inflammation process and inflammation- related diseases.

• Biomolecules & Therapeutics
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• 제21권5호
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• pp.381-390
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• 2013

The purpose of this study was to examine whether ginsenoside Rg3 (GRg3) could improve learning and memory impairments and inflammatory reactions induced by injecting lipopolysaccharide (LPS) into the brains of rats. The effects of GRg3 on proinflammatory mediators in the hippocampus and the underlying mechanisms of these effects were also investigated. Injection of LPS into the lateral ventricle caused chronic inflammation and produced deficits in learning in a memory-impairment animal model. Daily administration of GRg3 (10, 20, and 50 mg/kg, i.p.) for 21 consecutive days markedly improved the LPS-induced learning and memory disabilities demonstrated on the step-through passive avoidance test and Morris water maze test. GRg3 administration significantly decreased expression of pro-inflammatory mediators such as tumor necrosis factor-${\alpha}$, interleukin-1${\beta}$, and cyclooxygenase-2 in the hippocampus, as assessed by reverse transcription-polymerase chain reaction analysis and immunohistochemistry. Together, these findings suggest that GRg3 significantly attenuated LPS-induced cognitive impairment by inhibiting the expression of pro-inflammatory mediators in the rat brain. These results suggest that GRg3 may be effective for preventing or slowing the development of neurological disorders, including Alzheimer's disease, by improving cognitive and memory functions due to its anti-inflammatory activity in the brain.

• Natural Product Sciences
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• 제25권4호
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• pp.326-333
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• 2019

The purpose of our study was to evaluate anti-AD potential of Cirsium maackii flowers. MeOH extract, CH₂Cl₂, EtOAc, and n-BuOH fraction of this flower notably inhibited BACE1 (IC₅₀ = 76.47 ± 1.66, 22.98 ± 1.45, 8.65 ± 0.63, and 72.47 ± 3.04 ㎍/mL, respectively). β-amyrenone (49.70 mg) (1), lupeol acetate (1.43 g) (2), lupeol (1.22 g) (3), lupenone (23.70 mg) (4), β-sitosterol (1.01 g) (6), and β-sitosterol glucoside (13.00 mg) (7) from CH₂Cl₂, apigenin (100.20 mg) (8), luteolin (19.00 mg) (9), apigenin 7-O-glucuronide methyl ester (21.30 mg) (14), and tracheloside (53.70 mg) (5) from EtOAc, apigenin 5-O-glucoside (11.00 mg) (10), luteolin 5-O-glucoside (11.00 mg) (11) and apigenin 7-O-glucuronide (91.00 mg) (12) from n-BuOH, and luteolin 7-O-glucuronide (22.00 mg) (13) from H₂O fraction were isolated. HPLC showed high levels of 8, 9 and 12 in MeOH extract (33.07 ± 0.07, 31. 44 ± 0.17 and 16.89 ± 0.33 mg/g, respectively), EtOAc (161.01 ± 1.78, 96.93 ± 0.34 and 73.38 ± 0.06 mg/g, respectively), and n-BuOH fraction (32.18 ± 0.33, 44.31 ± 0.32 and 105.94 ± 0.36 mg/g, respectively). Since, 3 and 9 are well-known BACE1 inhibitors, the anti-AD activity of C. maackii flower might be attributable to their presence.

• 대한한방내과학회지
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• 제24권1호
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• pp.75-83
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• 2003

Objectives : Peroxynitrite($ONOO^-$), formed from the reaction of $O2^-$ and NO, is a cytotoxic species that can oxidize several cellular components such as proteins, lipids and DNA. It has been involved in the aging process and age-related diseases such as Alzheimer's disease, rheumatoid arthritis, cancer and atherosclerosis. The aim of this study was to investigate scavenging activities of $ONOO^-$ and its precursors, NO and $O_{2^-}$ and its scavenging mechanism of Zingiberis Rhizoma (ZR). Methods : To investigate scavenging activities of $ONOO^-,\;NO,\;O_{2^-}$ and its scavenging mechanism, we used fluorescent probes like DCFDA, DAF-2 and DHR 123. The $ONOO^-$ scavenging activity on ZR was assayed by measuring oxidized dihydrorhodamine 123 (DHR 123) by fluorometry. The scavenging efficacy was expressed as IC50, showing the concentration of each sample that is required to cause 50% inhibition of DHR 123 oxidation. In a separate study, the protective effect of ZR on $ONOO^-$-induced nitration of bovine serum albumin was investigated through immuno-assay with a monoclonal anti-nitryrosine antibody, and a horseradish peroxidase-conjugated anti-mouse secondary antibody from sheep. Results : ZR markedly scavenged authentic $ONOO^-,\;O_{2^-}$ and NO. It also inhibited $ONOO^-$ induced by $O_{2^-}$ and NO which are derived from SIN-1. The data demonstrated that ZR led to decreased $ONOO^-$ mediated nitration of tyrosine through electron donation. It also inhibited the nitration of bovine serum albumin induced by $ONOO^-$ in a dose-dependent manner. Furtheremore, it blocked LPS-induced ROS and RNS generation. Conclusions : These results suggest that ZR can be developed as an effective $ONOO^-$ scavenger for the prevention of aging process and age-related diseases.

• Journal of Applied Biological Chemistry
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• 제63권3호
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• pp.283-290
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• 2020

Oxidative stress is one of the contributors of neurodegenerative disorders including Alzheimer's disease. According to previous studies, Aster yomena (Kitam.) Honda (AY) possesses variable pharmacological activities including anti-coagulant and anti-obesity effect. In this study, we aimed to determine the neuroprotective effect of ethyl acetate fraction from Aster yomena (Kitam.) Honda (EFAY) against oxidative stress. Therefore, we carried out 3-(4,5-dimethylthiazol-2-yl)-2,3-diphenyl tetrazolium bromide, lactate dehydrogenase (LDH), and 2',7'-dichlorofluorescin diacetate assays in SH-SY5Y neuronal cells treated with hydrogen peroxide (H₂O₂). H₂O₂-treated control cells exhibited reduced viability of cells, and increased LDH release and reactive oxygen species (ROS) production compared to normal cells. However, treatment with EFAY restored the cell viability and inhibited LDH release and ROS production. To investigate the underlying mechanisms by which EFAY attenuated neuronal oxidative damage, we measured protein expressions using Western blot analysis. Consequently, it was observed that EFAY down-regulated cyclooxygenase-2 and interleukin-1β protein expressions in H₂O₂-treated SH-SY5Y cells that mediated inflammatory reaction. In addition, apoptosis-related proteins including B-cell lymphoma-2-associated X protein/B-cell lymphoma-2 ratio, cleaved caspase-9, and cleaved-poly (ADP-ribose) polymerase protein expressions were suppressed when H₂O₂-treated cells were exposed to EFAY. Our results indicate that EFAY ameliorated H₂O₂-induced neuronal damage by regulating inflammation and apoptosis. Altogether, AY could be potential therapeutic agent for neurodegenerative diseases.

• 생명과학회지
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• 제23권6호
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• pp.812-824
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• 2013

Reserpine은 항고혈압제로서 알츠하이머질병의 증상을 나타내는 Caenorhabditis elegans에서 세포독성을 감소시켜 마비를 억제하고 수명을 연장시키는 것으로 알려져 있다. 본 연구에서는 이러한 reserpine의 효능을 포유동물에서 확인하기 위하여, 알츠하이머질병의 병리적 특성과 연관된 주요인자의 변화를 30일 동안 reserpine을 투여한 Tg2576 마우스에서 관찰하였다. 그 결과, 공격행동(aggressive behavior)은 vehicle 투여 그룹에 비하여 reserpine 투여 그룹에서 유의적으로 감소하였으나 사회적 접촉(social contact)은 유의적인 변화가 없었다. 뇌의 해마부분에서 알츠하이머질병의 원인 중 하나인 $A{\beta}$-42의 축적은 reserpine 투여 그룹에서 유의적으로 감소하였고, $A{\beta}$-42의 농도도 대조군에 비하여 reserpine 투여 그룹에서 감소하였다. 더불어, ${\gamma}$-secretase의 구성단백질 중에서 PS-2, Pen-2, APH-1의 발현은 대조군에 비하여 reserpine 투여군에서 유의적으로 감소하였으나 NCT 발현은 변화가 없었다. 혈청에서 NGF의 농도는 Tg2576 마우스에서 감소하였다가 reserpine 투여한 그룹에서 유의적으로 증가하였으며, high affinity receptor의 신호전달과정에 포함된 단백질 중에서 reserpine 투여 그룹은 TrkA의 인산화가 증가하고 ERK 인산화는 감소되었다. 한편 low affinity receptor의 신호전달과정에서, $p75^{NTR}$과 Bcl-2의 발현은 vehicle 그룹에 비하여 reserpine 투여 그룹에서 유의적으로 발현이 증가하였으나 RhoA의 발현은 reserpine 투여 그룹에서 감소하였다. 따라서 이러한 결과는 reserpine은 포유동물 치매모델인 Tg2576 마우스에서 행동학적 변화, $A{\beta}$-42의 축적, NGF의 농도, NGF신호전달의 변화 등을 유도하며, 향후 치매치료제로서 가능성을 제시하고 있다.

• Biomolecules & Therapeutics
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• 제19권3호
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• pp.288-295
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• 2011

Amyloid beta ($A{\beta}$)-induced neurotoxicity is a major pathological mechanism of Alzheimer's disease (AD). In this study, we investigated the inhibitory effect of L-theanine, a component of green tea (Camellia sinensis) on $A{\beta}_{1-42}$-induced neurotoxicity and oxidative damages of macromolecules. L-theanine inhibited $A{\beta}_{1-42}$-induced generation of reactive oxygen species, and activation of extracellular signal-regulated kinase and p38 mitogenic activated protein kinase as well as the activity of nuclear factor kappa-B. L-theanine also signifi cantly reduced oxidative protein and lipid damage, and elevated glutathione level. Consistent with the reduced neurotoxic signals, L-theanine (10-50 ${\mu}g$/ml) concomitantly attenuated $A{\beta}_{1-42}$ (5 ${\mu}M$)-induced neurotoxicity in SK-N-MC and SK-N-SH human neuroblastoma cells. These data indicate that L-theanine on $A{\beta}$-induced neurotoxicity prevented oxidative damages of neuronal cells, and may be useful in the prevention and treatment of neurodegenerative disease like AD.