• 치위생과학회지
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• 제19권3호
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• pp.198-204
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• 2019

Background: Oxidative stress is a known to be associated with in the pathogenesis of many inflammatory diseases, including periodontitis. Ursolic acid is a pentacyclic triterpenoid with has antimicrobial, antioxidative, and anticancer properties. However, the role of ursolic acid in the regulating of osteogenesis remains undetermined. This study was aimed to elucidate the crucial osteogenic effects of ursolic acid and its ability to inhibit oxidative stress by targeting the immediate early response 3 (IER3)/nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. Methods: Cell proliferation was determined using water-soluble tetrazolium salt assay, cell differentiation was evaluated by alkaline phosphatase (ALP) activity, and formation of calcium nodules was detected using alizarin red S stain. Generation of reactive oxygen species (ROS) was determined using by DCFH-DA fluorescence dye in hydrogen peroxide ($H_2O_2$)-treated MG-63 cells. Expression levels of IER3, Nrf2, and heme oxygenase-1 (HO-1) were analyzed using western blot analysis. Results: Our results showed that ursolic acid up-regulated the proliferation of osteoblasts without any cytotoxic effects, and promoted ALP activity and mineralization. $H_2O_2$-induced ROS generation was found to be significantly inhibited on treatment with ursolic acid. Furthermore, in $H_2O_2$-treated cells, the expression of the early response genes: IER3, Nrf2, and Nrf2-related phase II enzyme (HO-1) was enhanced in the presence of ursolic acid. Conclusion: The key findings of the present study elucidate the protective effects of ursolic acid against oxidative stress conditions in osteoblasts via the IER3/Nrf2 pathway. Thus, ursolic acid may be developed as a preventative and therapeutic agent for mineral homeostasis and inflammatory diseases caused due to oxidative injury.

• Anatomy and Cell Biology
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• 제50권3호
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• pp.207-213
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• 2017

Glycogen synthase kinase $(GSK)-3{\beta}$ and related enzymes are associated with various forms of neuroinflammation, including spinal cord injury (SCI). Our aim was to evaluate whether lithium, a non-selective inhibitor of $GSK-3{\beta}$, ameliorated SCI progression, and also to analyze whether lithium affected the expression levels of two representative $GSK-3{\beta}$-associated molecules, nuclear factor erythroid 2-related factor-2 (Nrf-2) and heme oxygenase-1 (HO-1) (a target gene of Nrf-2). Intraperitoneal lithium chloride (80 mg/kg/day for 3 days) significantly improved locomotor function at 8 days post-injury (DPI); this was maintained until 14 DPI (P<0.05). Western blotting showed significantly increased phosphorylation of $GSK-3{\beta}$ (Ser9), Nrf-2, and the Nrf-2 target HO-1 in the spinal cords of lithium-treated animals. Fewer neuropathological changes (e.g., hemorrhage, inflammatory cell infiltration, and tissue loss) were observed in the spinal cords of the lithium-treated group compared with the vehicle-treated group. Microglial activation (evaluated by measuring the immunoreactivity of ionized calcium-binding protein-1) was also significantly reduced in the lithium-treated group. These findings suggest that $GSK-3{\beta}$ becomes activated after SCI, and that a non-specific enzyme inhibitor, lithium, ameliorates rat SCI by increasing phosphorylation of $GSK-3{\beta}$ and the associated molecules Nrf-2 and HO-1.

• Biomolecules & Therapeutics
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• 제29권1호
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• pp.90-97
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• 2021

Ultraviolet B (UVB) radiation causes DNA base modifications. One of these changes leads to the generation of 8-oxoguanine (8-oxoG) due to oxidative stress. In human skin, this modification may induce sunburn, inflammation, and aging and may ultimately result in cancer. We investigated whether phloroglucinol (1,3,5-trihydroxybenzene), by enhancing the expression and activity of 8-oxoG DNA glycosylase 1 (Ogg1), had an effect on the capacity of UVB-exposed human HaCaT keratinocytes to repair oxidative DNA damage. Here, the effects of phloroglucinol were investigated using a luciferase activity assay, reverse transcription-polymerase chain reactions, western blot analysis, and a chromatin immunoprecipitation assay. Phloroglucinol restored Ogg1 activity and decreased the formation of 8-oxoG in UVB-exposed cells. Moreover, phloroglucinol increased Ogg1 transcription and protein expression, counteracting the UVB-induced reduction in Ogg1 levels. Phloroglucinol also enhanced the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) as well as Nrf2 binding to an antioxidant response element located in the Ogg1 gene promoter. UVB exposure inhibited the phosphorylation of protein kinase B (PKB or Akt) and extracellular signal-regulated kinase (Erk), two major enzymes involved in cell protection against oxidative stress, regulating the activity of Nrf2. Akt and Erk phosphorylation was restored by phloroglucinol in the UVB-exposed keratinocytes. These results indicated that phloroglucinol attenuated UVB-induced 8-oxoG formation in keratinocytes via an Akt/Erk-dependent, Nrf2/Ogg1-mediated signaling pathway.

• Archives of Plastic Surgery
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• 제41권6호
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• pp.654-660
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• 2014

Background Reactive oxygen species (ROS) damages cell molecules, and modifies cell signaling. The nuclear factor E2-related factor (Nrf2) is a critical transcription regulator, which protects cells against oxidative damage. Nrf2 expression is increased in a large number of cancers. However, little information has been reported regarding the expression of Nrf2 in skin cancers. Hence, we explored the expression of Nrf2 protein in skin cancers. Methods The Nrf2 protein expression in 24 specimens, including 6 malignant melanomas (MM), 6 squamous cell carcinomas (SCC), 6 basal cell carcinomas (BCC), and 6 normal skin tissues, was evaluated by western blotting. Immunohistochemical staining was performed. The expression of Kelch-like ECH-associated protein 1 (Keap1), the key regulator of Nrf2, was also analyzed by western blotting. Results Small interfering RNA transfection to the melanoma cell line G361 confirmed that an approximately 66 kDa band was the true Nrf2 band. The western blot revealed that the Nrf2 protein was definitely expressed in normal skin tissues, but the Nrf2 expression was decreased in MM, SCC, and BCC. Immunohistochemical examination showed that expression of Nrf2 was decreased in all skin cancer tissues compared to the normal skin tissues. Keap1 was not expressed in all malignant skin tumors and normal skin tissues by western blot. Conclusions ROS was increased in various types of cancers which proteins were highly expressed or underexpressed. This study demonstrated that the expression of Nrf2 protein was down-regulated in human malignant skin tumors. We suggest that decreased expression of Nrf2 is related to skin cancers.

• The Korean Journal of Pain
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• 제34권1호
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• pp.35-46
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• 2021

Background: The present investigation explored the therapeutic actions of oleuropein along with the possible signaling pathway involved in attenuating neuropathic pain in chronic constriction injury (CCI) and vincristine-induced neuropathic pain in male rats. Methods: Four loose ligatures were placed around the sciatic nerve to induce CCI, and vincristine (50 ㎍/kg) was injected for 10 days to develop neuropathic pain. The development of cold allodynia, mechanical allodynia, and mechanical hyperalgesia was assessed using different pain-related behavioral tests. The levels of H2S, cystathionine-γ-lyase (CSE), cystathionine-β-synthase (CBS), orexin, and nuclear factor erythroid-2-related factor 2 (Nrf2) were measured in the sciatic nerve. Results: Treatment with oleuropein for 14 days led to significant amelioration of behavioral manifestations of neuropathic pain in two pain models. Moreover, oleuropein restored both CCI and vincristine-induced decreases in H2S, CSE, CBS, orexin, and Nrf2 levels. Co-administration of suvorexant, an orexin receptor antagonist, significantly counteracted the pain-attenuating actions of oleuropein and Nrf2 levels without modulating H2S, CSE and CBS. Conclusions: Oleuropein has therapeutic potential to attenuate the pain manifestations in CCI and vincristine-induced neuropathic pain, possibly by restoring the CSE, CBS, and H2S, which may subsequently increase the expression of orexin and Nrf2 to ameliorate behavioral manifestations of pain.

• 대한한의학방제학회지
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• 제26권1호
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• pp.1-12
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• 2018

Objectives : Cheongnoimyungshin-hwan (CNMSH) is a Herbal compound prescription that is composed mainly of herbal medicines such as Ginseng Radix Alba, Angelicae Gigantis Radix, Dioscoreae Rhizoma, Longan Arillus and cornus cervi parvum, and for the purpose of improving memory and preventing dementia. Methods : In this study, it was investigated whether CNMSH could suppress inflammatory response and oxidative stress in the lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophage cells. As a result, CNMSH decreased expression of inducible nitric oxide (NO) synthase and cyclooxygenase-2, and also inhibited production of NO, prostaglandin E2. Results : This effect was associated with the suppression of the expression of p65, one of the nuclear factor-kappaB ($NF-{\kappa}B$) subunits, and increased expression of $I{\kappa}B-{\alpha}$, inhibit the $NF-{\kappa}B$ transcription factor. In addition, CNMSH significantly blocked intracellular reactive oxygen species accumulation in response to LPS stimulation. Furthermore, CNMSH increased expression of nuclear factor erythroid 2-related factor (Nrf)-2 activation and heme oxygenase (HO)-1. Conclusions : Therefore, it has been shown anti-inflammatory and antioxidant effects by inhibiting the expression and production of inflammatory mediators in LPS-stimulated macrophages, and is associated with ROS generation and is activated by Nrf2/HO-1 signaling pathway.

• Journal of Microbiology and Biotechnology
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• 제20권9호
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• pp.1331-1338
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• 2010

This study was designed to investigate the potentially protective effects of Curcuma longa Linn. extract (CLE) on carbon tetrachloride ($CCl_4$)-induced hepatotoxicity in rats. Male Sprague-Dawley rats were pretreated with 50 or 100mg/kg of CLE or 100mg/kg of butylated hydroxytoluene(BHT) for 14 days before $CCl_4$ administration. In addition, the CLE control group was pretreated with 100mg/kg CLE for only 14 days. Three hours after the final treatment, a single dose of $CCl_4$ (20mg/kg) was administrated intraperitoneally to each group. After the completion of this phase of the experiment, food and water were removed 12 h prior to the next step. The rats were then anesthetized by urethane and their blood and liver were collected. It was observed that the aspartate aminotransferase and alanine aminotransferase activities of the serum, and the hepatic malondialdehyde levels had significantly decreased in the CLE group when compared with the $CCl_4$-treated group. The antioxidant activities, such as superoxide dismutase, catalase, and glutathione peroxidase activities, in addition to glutathione content, had increased considerably in the CLE group compared with the $CCl_4$-treated group. Phase II detoxifying enzymes, such as glutathione S-transferase, were found to have significantly increased in the CLE group as opposed to the $CCl_4$-treated group. The content of Nrf2 was determined by Western blot analysis. Pretreated CLE increased the level of nuclear translocated Nrf2, and the Nrf2 then increased the activity of the antioxidant and phase II detoxifying enzymes. These results indicate that CLE has protective effects against $CCl_4$-induced hepatotoxicity in rats, via activities of antioxidant and phase II detoxifying enzymes, and through the activation of nuclear translocated Nrf2.

• 한국자원식물학회지
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• 제35권5호
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• pp.565-573
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• 2022

치주조직에 존재하는 주요한 세포의 한 형태인 인체 치은섬유아세포는 다양한 구강유해세균으로부터 염증이 유발되어지며, 그중 대표적으로 치주염 원인균인 P. gingivalis의 내독소인 LPS-PG로부터 염증성 자극에 반응하여 다양한 염증매개 물질을 분비한다. 본 연구에서는 치주염을 일으키는 주요한 원인균 중 하나인 P. gingivalis로 부터 분리한 LPS-PG를 이용하여 인체 치은섬유아세포주인 HGF-1 세포에 염증을 유도한 후 LRE에 대한 항염증 및 항산화 효과를 분석하였다. 실험 결과, LRE는 LPS-PG 유도에 따라 iNOS에 의한 NO 생성과 COX-2에 의한 PGE₂와 같은 염증 매개 인자의 발현 및 생성 억제와 함께 염증성 싸이토카인(TNF-α, IL-1β및 IL-6)의 생성 또한 억제하였다. 신호전달계에서 염증성 전사인자의 발현 경로를 확인하기 위하여 TLR4/Myd88/NF-κB의 활성을 확인한 결과, LRE 처리에 따라 농도 의존적으로 억제되는 것을 확인하였다. 또한 산화 환원 효소로 항염증효과를 나타내는 것으로 알려진2상 효소 중 하나인 NQO-1과 이의 전사인자인 Nrf2를 분석 한 결과 LRE 처리에 의해 효소의 활성이 높아지는 것을 확인할 수 있었다. 결론적으로 LRE는 TLR4/Myd88/NF-κB 신호전달 경로를 억제하고 NQO1/Nrf2 활성을 유도함으로써 HGF-1 세포에서 LPS-PG에 의해 유도된 염증을 억제하는 것으로 사료되며, 향후 LRE는 식·의약품 소재 개발에서 치주질환 개선의 가능성이 있는 후보물질이 될 수 있을 것으로 사료된다.