유도된 산화적 스트레스 매개 세포사멸에 대한 baicalein 의 보호효과를
평가하였다. Baicalein 은 자외선 B 의 파장 범위 내의 빛을 흡수했다. 또한,
baicalein 은 자외선 B 의 조사에 반응하여 세포 내 활성산소종 (ROS) 의 수준을 감소시켰다. Baicalein 은 HaCaT 세포에서 자외선 B 의 조사에 유도된 DNA 손상,
Departments of 1 Advanced Convergence Technology & Science, 2 Veterinary Medicine, 3 Marine Life Sciences,
4 Civil and Enviromental Engineering, and 5 Nuclear and Energy Engineering, Jeju National University, Jeju 64243, Korea
(Received: January 22, 2016; Revised: March 13, 2016; Accepted: March 18, 2016)
Abstract : When exposed to gamma-rays, hair follicular cells immediately go through apoptosis, which hampers their rapid differentiation essential for the regeneration of hair. Phloroglucinol (PG) is a phenolic compound of Ecklonia cava, brown algae abundant in Jeju island, Korea. Containing plentiful polyphenols, PG is known for its instructive effects by inhibiting apoptosis, scavenging oxygen radicals, and protecting cells againstoxidativestress. In this study, we demonstrate that PG rescues radiosensitive hair follicular cells from gamma radiation-induced apoptosis and DNA damage. To identify protective capacity of PG on hair follicles, we irradiated with 8.5 Gy (1.5 Gy/min) of gamma- rays to the whole body of C57BL/6 mice at day 6 after depilation with or without PG. In mice exposed to radiation, the expression of proapoptotic molecule p53 was downregulated in the skin of PG treated group. On immunohistochemical observation of the skin, PG inhibited the immunoreactivity of p53 and cleaved caspase-3. PG treatment protected hair follicular cells from cell death due to gamma-radiation. Our results suggest that PG presents radioprotective effects by inhibiting apoptosis of radiosensitive hair follicular cells and can protect hair follicular cells from gamma-ray induced damage.
radical 소거능을 나타내어 본 연구와 유사한 수치를 나타냄을 알 수 있었다[27].
페놀성 화합물은 식물계에 널리 분포되어 있는 2차 대사산물 의 하나로써, 분자 내 phenolic hydroxyl기가 효소 단백질과 같 은 분자들과 결합하는 성질로 인해 항암, 항염증 및 항산화 등 의 다양한 생리활성 기능을 갖는 것으로 알려져 있다[28]. 플라 보노이드는 식물이 함유하고 있는 노란색을 띤 천연 물질을 총 칭하며, 항산화, 항균, 면역증강, 모세혈관 강화 등 다양한 생리 활성을 나타낸다[29]. 특히 페놀 및 플라보노이드는 우수한 항 산화 활성을 나타내며, 이들 함량이 높을수록 항산화 활성이 높 다고 알려져 있다[29,30]. 본 연구에서 G. uralensis, G. glabra, Table 1 DPPH radical scavenging activity of Glycyrrhiza uralensis,
group (catechol structure), enhance and play an important role in antiradical efficacy
of simple coumarins [26] . These results are consistent with a previous report that the
radical scavenging activity was higher for caffeic acid bearing o-dihydroxy group
than methoxy-substituted derivatives [27] . Thus it can be inferred that the resonance
H 2 O 2 treatment increased the levels of intracellular ROS and DNA double-strand breaks (DSBs) and decreased the level of Ku70 protein and the phosphorylation (activation) of DNA-dependent protein kinase catalytic subunit (DNA-PKcs), which are involved in the repair of DSBs by nonhomologous end joining. Baicalein effectively scavenged intracellular ROS induced by H 2 O 2 , reduced DSBs, and rescued Ku70 protein level and phosphorylation of DNA-PKcs. In cellular response to DNA base damage, 8-oxoguanine DNA glycosylase 1 (OGG1) plays a vital role in the removal of 8-oxoguanine (8-OxoG), which is formed mainly by oxidativestress. Baicalein significantly decreased the levels of 8-OxoG induced by H 2 O 2 , and this correlated with increases in OGG1 promoter activity and OGG1 mRNA and protein expression. The phosphorylated form of Akt kinase, which is a regulator of OGG1, was sharply decreased by H 2 O 2 , but was prevented by baicalein. A specific Akt inhibitor abolished the cytoprotective effects ofbaicalein, suggesting that OGG1 induction by baicalein involves the Akt pathway. In conclusion, baicalein exerted protective effects against DNA damage induced by oxidativestress by activating DNA repair systems and scavenging ROS.
In other studies, administration of anesthetic agents was found to result in pharmacologic preconditioning againstoxidative injury [20,21].
Autophagy is a conserved cellular catabolic process that can engulf cytoplasmic components and degrade them through the lysosomal pathway, thus, they can then be recycled. Generally, autophagy is thought to be induced under stress conditions [22-24]. However, whether the induction of autophagy contributes to cell survival or cell death remains elusive. Oxidative injury is one of the most common causes of bone remodeling inhibition. Therefore, a sustainable drug is needed to identify better, safer anabolic agents with low cytotoxicity that act by either increasing osteoblast proliferation or differentiation to enhance bone formation [25]. Several lines of evidence have shown that anesthesia drugs may have a beneficial effect on bone loss and fracture outcomes [26]. On the basis of this, it was hypothesized that the antioxidant properties of propofol may protect human fetal osteoblast (hFOB) cells againstoxidative injury caused by H 2 O 2 . Therefore, the aim of this study was to evaluate the protective effects of propofol- mediated autophagy activation on oxidativestressinduced by H 2 O 2 treatment in the hFOB cell line.
However, pretreatment with 3-MA inhibited LC3-II expression (Fig. 5).
DISCUSSION
In this study, the protectiveeffectof PPC againstoxidative-stress-induced COS-7 cell apoptosis was pro- ven in an in vitro experimental model of H 2 O 2 -inducedoxidative injury. We also demonstrated that the anti- oxidant effectof PPC was mediated by cellular autophagy activation by showing the formation of autophagosomes in COS-7 cells by using a fluorescence microscope and the expression of autophagy-related proteins by using western blot analysis.
Microglial apoptosis 개선 효과
뇌에서 대식세포의 역할을 하는 microglia인 BV-2 세포는 신경 계의 면역세포로 작용하고, BV-2 세포는 외부 및 내부 자극을 통 해 활성화되어 다양한 전 염증성 사이토카인을 분비한다. 외부 및 내부 자극으로 인한 염증이 과도하게 발생하게 되면, 신경 퇴 화에 관여하는 신경 염증을 유발하게 된다(Bozic 등, 2015). 따라 서 외부 자극으로부터 염증반응을 매개하기 위해 세포막에 존재 하는 Toll-like receptor (TLR) 복합체가 활성화되고, 이는 신호 전 달에 의해 세포 증식 및 생존에 관여하는 phosphatidylinositol-3- kinase (PI3K)/Akt 경로와 세포 자살에 관여하는 c-Jun N-terminal kinases (JNK) 경로 및 nuclear factor kappa light chain enhancer of activated B cells (NF-κB) 경로 등이 조절된다(Khanna 등, 2015). 하지만 과도한 산화적 스트레스는 염증반응으로 인한 세 포 자살의 경로를 활성화시키고, 이들은 Akt, JNK, NF-κB 등을 포함하는 전사인자의 인산화를 유도한다. 또한, TNF-α, IL-1β 및 IL-6 와 같은 염증성 사이토카인을 과분비하며, caspase-3, caspase- 7 과 같은 인자의 발현을 유도해 caspase 의존성 세포 사멸을 유 발한다(Ni 등, 2015; Reuter 등, 2010). PM 2.5 는 이러한 면역계 세 포를 활성화시켜 염증반응을 매개할 수 있으며, BV-2 세포의 활 성화는 뇌 신경염증을 유도해 결과적으로 뇌 신경세포의 손상을 초래한다. 따라서 본 연구에서는 PM 2.5 를 처리한 BV-2 세포에서 세포 사멸 및 염증 반응에 관여하는 인자의 발현 수준을 확인함 으로써 EFEL이 가지는 개선 효과를 확인하였다.
Bcl-2 ratio, whereas EFAY treatment down-regulated Bax/Bcl-2 ratio. Furthermore, up-regulated protein expressions of cleaved caspase-9 and cleaved PARP were observed subsequent to treatment of SH-SY5Y cells with H 2 O 2 . However, treatment with EFAY also down-regulated cleaved caspase-9 and cleaved PARP expressions, suggesting that EFAY exerts a protectiveeffectagainst apoptosis in H 2 O 2 -treated neuronal cells. In addition, isoquercitrin, one of the active compounds of EFAY acts as an antioxidant and regulates apoptosis-related protein expressions of Bcl-2 and cleaved PARP in H 2 O 2 -treated RGC-5 cells [55]. Isoquercitrin was also reported to inhibit H 2 O 2 -induced apoptosis of EA.hy926 cells by mediating the PI3K/Akt/GSK3β signaling [56]. These results indicated that EFAY exerts a protectiveeffect on the apoptotic effectagainst H 2 O 2 -induced neuronal damage in SH- SY5Y cells by down-regulating Bax/Bcl-2 ratio and caspase cascades.
- 31 -
DISCUSSION
Oxidative stress plays a pivotal role in cellular injury induced by hyperglycemia. High glucose level can stimulate free radical production. Thus, a weak defense system becomes unable to counteract the enhanced ROS generation and, as a result, an imbalance occurs which leads to oxidativestress [20, 21]. In the present study, we demonstrated that 7,8-DHF dramatically inhibited high glucose-induced cell death, apoptosis, and mitochondrial dysfunction. 7,8-DHF protectiveeffect might be attributed to its powerful antioxidant action, as evidenced by the markedly reduced ROS.
In addition, delphinidin, the active compound from eggplant, exerted the strong 1,1-diphenyl-2-picrylhydrazyl scavenging effect with IC 50 value of 6.59 µg/mL. Furthermore, the cellular oxidativestress was induced by 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH) in LLC-PK 1 cells, while treatment of delphinidin atteunated AAPH-inducedoxidativestress as dose-dependent manner. The present study suggests the antioxidative activity of eggplant and delphinidin against free radical-inducedoxidativestress.
Abstract : To increase antioxidative activity of Chungkukjang, the protective effect of Seoritae Chungkukjang (SC) added with green tea powder against oxidative stress was evaluated un[r]
Microglial apoptosis 개선 효과
뇌에서 대식세포의 역할을 하는 microglia인 BV-2 세포는 신경 계의 면역세포로 작용하고, BV-2 세포는 외부 및 내부 자극을 통 해 활성화되어 다양한 전 염증성 사이토카인을 분비한다. 외부 및 내부 자극으로 인한 염증이 과도하게 발생하게 되면, 신경 퇴 화에 관여하는 신경 염증을 유발하게 된다(Bozic 등, 2015). 따라 서 외부 자극으로부터 염증반응을 매개하기 위해 세포막에 존재 하는 Toll-like receptor (TLR) 복합체가 활성화되고, 이는 신호 전 달에 의해 세포 증식 및 생존에 관여하는 phosphatidylinositol-3- kinase (PI3K)/Akt 경로와 세포 자살에 관여하는 c-Jun N-terminal kinases (JNK) 경로 및 nuclear factor kappa light chain enhancer of activated B cells (NF-κB) 경로 등이 조절된다(Khanna 등, 2015). 하지만 과도한 산화적 스트레스는 염증반응으로 인한 세 포 자살의 경로를 활성화시키고, 이들은 Akt, JNK, NF-κB 등을 포함하는 전사인자의 인산화를 유도한다. 또한, TNF-α, IL-1β 및 IL-6 와 같은 염증성 사이토카인을 과분비하며, caspase-3, caspase- 7 과 같은 인자의 발현을 유도해 caspase 의존성 세포 사멸을 유 발한다(Ni 등, 2015; Reuter 등, 2010). PM 2.5 는 이러한 면역계 세 포를 활성화시켜 염증반응을 매개할 수 있으며, BV-2 세포의 활 성화는 뇌 신경염증을 유도해 결과적으로 뇌 신경세포의 손상을 초래한다. 따라서 본 연구에서는 PM 2.5 를 처리한 BV-2 세포에서 세포 사멸 및 염증 반응에 관여하는 인자의 발현 수준을 확인함 으로써 EFEL이 가지는 개선 효과를 확인하였다.
Hyung Don Kim, Eun Young Lee, Jeong-Yong Park, Kyung Hye Seo, Kang-Hyo Lee, Jehun Choi, Jae-Gu Han, Jae-Han Cho*, and Seung Eun Lee*
Department of Herbal Crop Research, NIHHS, RDA, Eumseong, Chungbuk 27709, Korea.
ABSTRACT: This study was carried out to identify medicinal mushrooms with protective effects againstoxidativestress in PC12 neuronal cell line, followed by evaluation of their antioxidant property. Extracts of medicinal mushrooms, including Ganoderma lucidum extract (GLE), antler-shaped Ganoderma lingzhi extract (AGLE), Hericium erinaceus extract (HEE), and Sanghuangporus baumii extract (SBE), were screened for cytotoxicity using MTT assay. None of the extracts up to 10 µg/ml concentration affected cell viability. These extracts were further checked for their protectiveeffectagainstoxidativestress-induced reactive oxygen species (ROS) production. Exposure to 50 µM H 2 O 2 induced ROS generation in PC12 cells, which was inhibited only by treatment with AGLE. In addition, inhibition of H 2 O 2 -induced ROS generation by AGLE was found to be in a dose-dependent manner (2.5, 5, and 10 µg/ml). Microscopic examination of DCF fluorescence for detection of ROS showed a similar pattern.
Effect of the EtOAc fraction of PT on LDH release in H 2 O 2 - treated SH-SY5Y cells
LDH is an enzyme present in the cytoplasm of most tissues including the brain. The organ and extracellular levels of LDH are used as an indicator of cell damage [40]. A previous study demonstrated that H 2 O 2 exposure increased the LDH release induced by oxidative damage in SH-SY5Y cells [41]. To confirm the neuroprotective effect on oxidativestress, we investigated the effectof the EtOAc fraction of PT on LDH release in H 2 O 2 -treated SH-SY5Y cells (Fig. 3). The H 2 O 2 -treated control group had significantly increased LDH release (100%) compared to the non- treated normal group of 85.13%. Treatment with the EtOAc fraction of PT at doses of 1, 5, and 10 µg/mL significantly decreased the LDH release compared to the H 2 O 2 -treated control group. Therefore, we suggested that PT prevented the neuronal damage caused by oxidativestress by decreasing LDH release.
Cell viability(%) = 100×AT/AC, AC; absorbance of control, AT; absorbance of tested extract solution
4) Hydrogen peroxide-induced cytotoxicity 측정 Hydrogen peroxide에 의해 유도된 cytotoxicity 에 대한 보호효과를 알아보기 위해 Mosmann 등 9) 의 MTT test를 응용하여 다음과 같이 실험하였 다. 96 well plate에 1×10 5 cells/ml의 cell을 100 μ l씩 넣고 37℃, 5% CO 2 incubator에서 24 시간동 안 배양한 후 배지를 버리고 배양세포 표면을 PBS 용액으로 씻어주었다. PBS에 녹인 각각의 시료 1, 5, 10, 50, 100 μg/ml과 FBS free DMEM 에 녹인 50 μM H O 을 각각의 well에 처리한
The protective effect of Perilla frutescens from ONOO - -induced oxidative stress and antiaging effect under cellular system.. Hyun Young Kim 1 , Bo Ra Hwang 2 , Ting Ting Wu 2 , Eun Ju[r]
Methods : Component analysis of Citri Unshius Pericarpium was analyzed by UPLC with C 18 column. Cell viability was determined by MTT assay. The enzyme activity of superoxide dismutase (SOD) and the level of reactive oxygen species (ROS) and reduced glutathione (GSH) were analyzed using commercially available kits.
Results : Cadmium caused severe HepG2 cell death. Cadmium also increased ROS production, consistent with depletion of GSH and inhibition of the SOD enzyme. However, CEE treatment reduced cell death and relieved oxidativestress caused by cadmium toxicity. CEE lowered ROS levels and improved depletion of GSH levels. CEE also enhanced the enzymatic activity of SOD. In component analysis, hesperidin was the most abundant of the five marker compounds (Narigenin, Narigin, Narirutin, Hesperidin and Hesperidin), which assumes that hesperidin partially contributed to the antioxidant activity of CEE.
요 약
본 연구에서는 다양한 만성 대사성 질환에 기초적 유발 기전 인 산화적 스트레스 및 염증에 대한 폴리페놀 함유 블랙 초크베 리의 보호 효과를 살펴보았다. 블랙 초크베리는 DPPH, ·OH, 및 NO 등 다양한 라디컬 소거능을 나타냈다. 내독소인 LPS에 의해 활성화된 대식세포 내 ROS 증가는 폴리페놀 함유 블랙 초크베 리에 의해 감소하였다. 이는 블랙 초크베리가 전사인자인 NRF2 의 핵 내 이동 및 이로 인해 조절되는 SOD에 의한 ROS 소거능 Fig. 2. Effects of CBE on singnaling pathway in RAW 264.7 macrophages. (A) Nuclear translocation of NF- κB (B) COX-2 and iNOS expression and (C) COX-2 and iNOS protein levels. Data are expressed as relative expression to control. Bars with different letters are significantly different (p<0.05). Values are means±SEM; n=6. GAPDH and TBP were used for the puritiy of cytoplasmic and nuclear fractions, respectively. CBE, polyphenol-rich chokeberry extract; COX-2, cyclooxygenase-2; GAPDH, glyceraldehyde 3-phosphate dehydrogenase;