Prostaglandin A₂-induced Apoptosis is Not Inhibited by Heme Oygenase-1 in U2OS Cells

Kyoung-Won Ko(고경원),Sun-Young Lee(이선영),Ji-Hyun Ahn(안지현),Jaetaek Kim(김재택),In-Kyung Kim(김인경),Ho-Shik Kim(김호식) 한국생명과학회 2008 생명과학회지 Vol.18 No.11

Prostaglandin A₂ (PGA₂)는 사람 골육종 세포인 U2OS 세포주에서 apoptosis와 heme oxygenase (HO)-1의 발현을 함께 유도하였다. PGA₂에 의한 apoptosis는 HO-1의 과도한 발현이나 HO-1에 대한 small interfering RNA에 의한 발현저하에 의하여 변동되지 않았으나 H₂O₂에 의한 세포사망은 HO-1의 발현 수준에 반비례하여 변동되었다. 또한 thiol antioxidant인 N-acetyl-L-cysteine (NAC)은 PGA₂에 의한 세포사망과 HO-1의 발현 증가를 모두 차단하였지만, non-thiol antioxidant인 butylated hydroxyanisole (BHA)과 ascorbic acid는 세포사망과 HO-1의 발현 유도를 차단하지 않았다. 이와 같은 결과들은 PGA₂는 산화성 손상에 의해서가 아니라 PGA₂의 thiol-reactivity에 의하여 apoptosis와 HO-1의 발현을 유도하며, HO-1의 발현은 PGA₂에 의한 apoptosis와는 독립적인 현상이거나 기능적으로 apoptosis 유도의 하부에 위치하고 apoptosis의 진행에는 기여하지 않을 것이라는 것을 시사해 준다. Prostaglandin A₂ (PGA₂), one of cyclopentenone PGs, induced both apoptosis and heme oxygenase (HO)-1 expression in U2OS cells. PGA₂-induced apoptosis was not perturbed by either over-expression or knock-down of HO-1, whereas H₂O₂-induced cell death was inversely modulated by the expression level of HO-1. In addition, N-acetyl-L-cysteine (NAC), a thiol antioxidant, blocked both apoptosis and HO-1 expression induced by PGA₂. But, non-thiol antioxidants like butylated hydorxyanisole (BHA) and ascorbic acid did not block either apoptosis or HO-1-induction. Taken together, these results suggest that PGA₂ induces both apoptosis and HO-1 expression, which are critically related to the thiol-reactivity of PGA₂, but not oxidative stress, and HO-1 expression may be independent or functionally located downstream of apoptosis by PGA₂ without contribution to apoptosis progression.

제주도 지하수 질산염 농도의 시·공간적 변화 특성: 장기(1993-2015) 모니터링 자료의 평가

김호림(Ho-Rim Kim),오준섭(Junseop Oh),도현권(Hyun-Kwon Do),이경진(Kyung-Jin Lee),현익현(Ik-Hyun Hyun),오상실(Sang-Sil Oh),감상규(Sang-Kyu Kam),윤성택(Seong-Taek Yun) 대한자원환경지질학회 2018 자원환경지질 Vol.51 No.1

1993년부터 2015년까지 관측된 제주도 지하수 장기모니터링 관측정(N = 4,835)에서 수집된 지하수 수질자료(N = 21,568)를 기반으로 질산성질소의 시공간적 변동 특성을 평가하였다. 제주도 지하수의 질산성질소 농도의 중앙값은 2.5 mg/L로서 다른 국가나 대륙의 조사 결과에 비해 다소 높거나 유사한 것으로 나타났다. 또한 지하수 용도, 행정구역 및 고도 별로 유의한 차이를 보였다. 특히, 산간 지역에 비해 저지대 해안가에 위치한 농업 및 주거지역에서 농도가 높음을 확인하였다. Mann-Kendall 및 Sen’s slope 분석을 활용한 질산성질소 농도의 추세 분석 결과, 하류 저지 대에 비해 중산간지역에서의 질산성질소 농도 증가 경향이 뚜렷하였다. 제주도 내 토지 피복의 시계열 변화 특성과 결부 지어 보면, 중산간지역의 오염 증가 추세는 농업지역의 확장 등 인위적 활동 증가에 기인한 결과로 판단된다. 반면,기지정된 지하수자원특별관리구역에서는 전반적으로 질산성질소 농도의 감소 경향이 나타났는데, 이는 지하수 관리 측면에서 수질관리를 위한 적극적인 정책이 유효함을 시사한다. 본 연구에서는 제주도 지하수의 질산성질소 오염관리를 위한 적정 방안을 제안한다. The spatio-temporal variations of nitrate concentrations in groundwater of Jeju Island were evaluated by an analysis of time series groundwater quality data (N = 21,568) that were collected from regional groundwater monitoring (number of wells = 4,835) for up to 20 years between 1993 and 2015. The median concentration of NO 3 -N is 2.5 mg/L, which is slightly higher than those reported from regional surveys in other countries. Nitrate concentrations of groundwater in wells tend to significantly vary according to different water usage (of the well), administrative districts, and topographic elevations: nitrate level is higher in low-lying agricultural and residential areas than those in high mountainous areas. The Mann-Kendall trend test and Sen’s slope analysis show that nitrate concentration in mid-mountainous areas tends to increase, possibly due to the expansion of agricultural areas toward highland. On the other hand, nitrate concentrations in the Specially Designated Groundwater Quality Protection Zones show the temporally decreasing trend, which implies the efficiency of groundwater management actions in Jeju. Proper measures for sustainable groundwater quality management are suggested in this study.

Combined Gene Therapy with Hypoxia-Inducible Factor-1α and Heme Oxygenase-1 for Therapeutic Angiogenesis

Bhang, Suk Ho,Kim, Ju Hee,Yang, Hee Seok,La, Wan-Geun,Lee, Tae-Jin,Kim, Ga Hee,Kim, Hyun Ah,Lee, Minhyung,Kim, Byung-Soo Mary Ann Liebert 2011 Tissue engineering. Part A Vol.17 No.7

<P>Transfection with either hypoxia-inducible factor-1α (HIF-1α) or heme oxygenase-1 (HO-1) gene can induce neovascularization in ischemic tissues. Although expression of transfected HIF-1α gene occurs rapidly, the expressed HIF-1α protein degrades quickly, limiting its therapeutic efficacy. Meanwhile, expressed HO-1 protein does not rapidly undergo degradation, but gene expression occurs a couple of days after transfection, resulting in apoptosis and a delay in angiogenesis in ischemic tissues at the incipient period of HO-1 gene transfection. We hypothesize that combined delivery of HIF-1α and HO-1 gene will enhance antiapoptosis and neovascularization in ischemic tissue compared with HIF-1α or HO-1 single-gene therapy. To test this hypothesis, ischemic mouse hindlimbs were treated with HIF-1α and/or HO-1 gene therapy. The combined gene therapy proved superior to both single-gene therapies, resulting in rapid expression of HIF-1α gene and long-term maintenance of expressed HO-1 protein. The apoptosis in the ischemic region was significantly less, and angiogenic growth factor secretion and angiogenesis were greater in the combined gene therapy than in either of the single-gene therapies. Our results suggest that a combined gene therapy of HIF-1α and HO-1 enhances the transfection of both genes and improves angiogenesis compared with either single-gene therapy.</P>

Induction of heme oxygenase-1 protects against podocyte apoptosis under diabetic conditions

Lee, Sang Choel,Han, Seung Hyeok,Li, Jin Ji,Lee, Sun Ha,Jung, Dong-Sub,Kwak, Seung-Jae,Kim, Seung Hye,Kim, Dong Ki,Yoo, Tae-Hyun,Kim, Jin Hyun,Chang, Se-Ho,Han, Dae Suk,Kang, Shin-Wook International Society of Nephrology 2009 Kidney international Vol.76 No.8

Heme oxygenase-1 (HO-1) is an anti-oxidant enzyme normally upregulated in response to oxidant injury. Here we determined the role of HO-1 in podocyte apoptosis in glomeruli of streptozotocin-treated rats and in immortalized mouse podocytes cultured in media containing normal or high glucose. HO-1 expression, its activity, the ratio of Bax/Bcl-2 protein, and active caspase-3 fragments were all significantly higher in isolated glomeruli of diabetic rats and in high glucose–treated podocytes. These increases were inhibited by zinc protoporphyrin treatment of the rats or by HO-1 siRNA treatment of the podocytes in culture. The number of apoptotic cells was also significantly increased in the glomeruli of diabetic rats and in high glucose–treated podocytes. Inhibition of HO-1 accentuated the increase in apoptotic cells both in vivo and in vitro. Our findings suggest that HO-1 expression protects against podocyte apoptosis under diabetic conditions.

15d-PGJ2 inhibits NF-kB and AP-1-mediated MMP-9 expression and invasion of breast cancer cell by means of a heme oxygenase-1-dependent mechanism

장혜연,On-Yu Hong,Hyun Jo Youn,김민걸,Cheorl-Ho Kim,정성후,Jong-Suk Kim 생화학분자생물학회 2020 BMB Reports Vol.53 No.4

Activation of peroxisome proliferator-activated receptor  (PPAR) serves as a key factor in the proliferation and invasion of breast cancer cells and is a potential therapeutic target for breast cancer. However, the mechanisms underlying this effect remain largely unknown. Heme oxygenase-1 (HO-1) is induced and overexpressed in various cancers and is associated with features of tumor aggressiveness. Recent studies have shown that HO-1 is a major downstream target of PPAR. In this study, we investigated the effects of induction of HO-1 by PPAR on TPAinduced MMP-9 expression and cell invasion using MCF-7 breast cancer cells. TPA treatment increased NF-B /AP-1 DNA binding as well as MMP-9 expression. These effects were significantly blocked by 15d-PGJ2, a natural PPAR ligand. 15d-PGJ2 induced HO-1 expression in a dose-dependent manner. Interestingly, HO-1 siRNA significantly attenuated the inhibition of TPA-induced MMP-9 protein expression and cell invasion by 15d-PGJ2. These results suggest that 15d-PGJ2 inhibits TPA-induced MMP- 9 expression and invasion of MCF-7 cells by means of a heme oxygenase-1-dependent mechanism. Therefore, PPAR/HO-1 signaling- pathway inhibition may be beneficial for prevention and treatment of breast cancer.

Morin exerts cytoprotective effects against oxidative stress in C2C12 myoblasts via the upregulation of Nrf2-dependent HO-1 expression and the activation of the ERK pathway

Lee, Moon Hee,Han, Min Ho,Lee, Dae-Sung,Park, Cheol,Hong, Su-Hyun,Kim, Gi-Young,Hong, Sang Hoon,Song, Kyoung Seob,Choi, Il-Whan,Cha, Hee-Jae,Choi, Yung Hyun UNKNOWN 2017 INTERNATIONAL JOURNAL OF MOLECULAR MEDICINE Vol.39 No.2

<P>In the present study, we investigated the cytoprotective efficacy of morin, a natural flavonoid, against oxidative stress and elucidated the underlying mechanisms in C2C12 myoblasts. Our results indicated that morin treatment prior to hydrogen peroxide (H2O2) exposure significantly increased cell viability and prevented the generation of reactive oxygen species. H2O2-induced comet-like DNA formation and gamma H2AX phosphorylation were also markedly suppressed by morin with a parallel inhibition of apoptosis in C2C12 myoblasts, suggesting that morin prevented H2O2-induced cellular DNA damage. Furthermore, morin markedly enhanced the expression of heme oxygenase-1 (HO-1) associated with the induction and phosphorylation of nuclear factor-erythroid 2-related factor 2 (Nrf2) and the inhibition of Kelch-like ECH-associated protein 1 (Keapl) expression. Notably, these events were eliminated by transient transfection with Nrf2-specific small interfering RNA. Additional experiments demonstrated that the activation of the Nrf2/HO-1 pathway by morin was mediated by the extracellular signal-regulated kinase (ERK) signaling cascade. This phenomenon was confirmed with suppressed Nrf2 phosphorylation and consequently diminished HO-1 expression in cells treated with a pharmacological inhibitor of ERK. Collectively, these results demonstrated that morin augments the cellular antioxidant defense capacity through the activation of Nrf2/HO-1 signaling, which involves the activation of the ERK pathway, thereby protecting C2C12 myoblasts from H2O2,-induced oxidative cytotoxicity.</P>

사례보고 : 하수오 복용 후 발생한 재발성 독성 간염 1예

배상훈 ( Sang Hoon Bae ),김동현 ( Dong Hyun Kim ),배영석 ( Young Seok Bae ),이광재 ( Kwang Jae Lee ),김동완 ( Dong Wan Kim ),윤정빈 ( Jeoung Bin Yoon ),홍준호 ( Joon Ho Hong ),김상현 ( Sang Hyun Kim ) 대한간학회 2010 Clinical and Molecular Hepatology(대한간학회지) Vol.16 No.2

Toxic hepatitis has been reported as a major cause of acute hepatitis, but its potential induction by herbal remedies and/or health foods is usually neglected. We experienced a case of toxic hepatitis associated with Polygoni multiflori, a Chinese herb commonly known as Ho-Shou-Wu. A 54-year-old woman consumed Ho-Shou-Wu for 1 month, after which she experienced fatigue and overall weakness. A diagnosis of toxic hepatitis was made based on her clinical history, the findings for viral markers and other laboratory data, and ultrasonography. Her condition improved considerably after she stopped taking Ho-Shou-Wu. However, she resumed taking Ho-Shou-Wu immediately after discharge from hospital, which aggravated her symptoms and liver function. She was immediately readmitted and stopped taking Ho-Shou-Wu. Her relapse into hepatitis immediate after resuming consumption of the herb is strongly indicative of the validity of Koch`s postulate in this case.

Ethanol Extract of Ganoderma lucidum Augments Cellular Anti-oxidant Defense through Activation of Nrf2/HO-1

Lee, Yoo-hwan,Kim, Jung-hee,Song, Choon-ho,Jang, Kyung-jeon,kim, Cheol-hong,Kang, Ji-Sook,Choi, Yung-hyun,Yoon, Hyun-Min KOREAN PHARMACOPUNCTURE INSTITUTE 2016 Journal of pharmacopuncture Vol.19 No.1

Objectives: The mushroom Ganoderma lucidum has been widely used as a traditional herbal medicine for many years. Although several studies have focused on the anti-oxidative activity of this mushroom, the molecular mechanisms underlying its activity have not yet been clearly established. The present study investigated the cytoprotective effect of ethanol extract of Ganoderma lucidum (EGL) against oxidative stress (hydrogen peroxide, $H_2O_2$) and elucidated the underlying mechanisms in a C2C12 myoblast cell line. Methods: Oxidative stress markers were determined by using the comet assay to measure reactive oxygen species (ROS) generation and deoxyribonucleic acid (DNA) damage. Cell viability and Western blotting analyses were employed to evaluate the cellular response to EGL and $H_2O_2$ in C2C12 cells. Transfection with nuclear factor erythroid 2-related factor 2 (Nrf2)-specific small interfering ribonucleic acid (siRNA) was conducted to understand the relationship between Nrf2 expression and $H_2O_2$-induced growth inhibition. Results: The results showed that EGL effectively inhibited $H_2O_2$-induced growth and the generation of ROS. EGL markedly suppressed $H_2O_2$-induced comet-like DNA formation and phosphorylation of histone H2AX at serine 139 ($p-{\gamma}H2AX$), a widely used marker of DNA damage, suggesting that EGL prevented $H_2O_2$-induced DNA damage. Furthermore, the EGL treatment effectively induced the expression of Nrf2, as well as heme oxygenase-1 (HO-1), with parallel phosphorylation and nuclear translocation of Nrf2 in the C2C12 myoblasts. However, zinc protoporphyrin IX, a HO-1 inhibitor, significantly abolished the protective effects of EGL against $H_2O_2$-induced accumulation of ROS and reduced cell growth. Notably, transient transfection with Nrf2-specific siRNA attenuated the cytoprotective effects and HO-1 induction by EGL, indicating that EGL induced the expression of HO-1 in an Nrf2-dependent manner. Conclusion: Collectively, these results demonstrate that EGL augments the cellular anti-oxidant defense capacity through activation of Nrf2/HO-1, thereby protecting C2C12 myoblasts from $H_2O_2$-induced oxidative cytotoxicity.

연잎, 연자육, 연자방 에탄올 추출물의 항염증 활성

이은주(Eun-Joo Lee),서유미(Yu-Mi Seo),김용현(Yong-Hyun Kim),정정욱(Chungwook Chung),성화정(Hwa-Jung Sung),손호용(Ho-Yong Sohn),박종이(Jong-Yi Park),김종식(Jong-Sik Kim) 한국생명과학회 2019 생명과학회지 Vol.29 No.4

연은 아시아 국가에서 음식과 약재로 널리 사용되는 재료이다. 본 연구에서는 연의 잎(leaf, NL), 자육(seed, NS), 자방(seedpod, NSP)으로부터 에탄올 추출물을 제조하고 이들의 항염증 활성과 작용기전을 규명하였다. 이들의 항염증 활성을 연구하기 위하여 LPS로 자극된 RAW 264.7 세포에서 nitric oxide (NO) 생산을 측정하였다. NL, NS, NSP는 세포 생존율에 영향을 주지 않으면서, 농도의존적으로 NO의 생산을 현저하게 저해하였으며, iNOS 및 COX-2와 같은 pro-inflammatory 중재자들의 단백질 발현을 감소시켰다. 또한, NL, NS, NSP는 MAPKs 단백질의 인신화를 감소시키고 NF-κB p65의 핵으로의 이동을 저해함으로써, 세 추출물에 의한 항염증 활성은 MAPKs 경로와 NF-κB 경로를 조절함으로써 이루어짐을 제시한다. 게다가, ROS의 생성이 세 추출물에 의해서 모두 저해되었으며, HO-1의 발현과 HO-1의 전사조절인자인 Nrf2의 핵으로의 이동이 증가되었다. 결론적으로, 이러한 연구결과는 연의 다양한 부위의 추출물인 NL, NS 그리고 NSP는 항염증 활성을 가지고 있으며, MAPKs, NF-κB, Nrf2/HO-1 등 다양한 신호경로를 통해 조절할 수 있음을 제시한다. Nelumbo nucifera, also known as sacred lotus, has mainly been used as a food throughout the Asian countries. In the present study, we prepared ethanol extracts from leaf (NL), seed (NS), and seedpod (NSP) of Nelumbo nucifera and investigated their anti-inflammatory activities in mouse macrophage RAW 264.7 cells. To evaluate the anti-inflammatory activities of NL, NS, and NSP, nitric oxide (NO) production was measured in LPS-stimulated RAW 264.7 cells. NL, NS, and NSP significantly reduced NO production in a dose-dependent manner without affecting cell viabilities. NL, NS, and NSP dramatically decreased the protein expression of pro-inflammatory genes such as iNOS and COX-2. NL, NS, and NSP also suppressed phosphorylation of MAPKs and the nuclear translocation of NF-κB p65 indicating they have their anti-inflammatory activities via regulating mitogen-activated protein kinases (MAPKs) and nuclear factor kappa B (NF-κB) pathways. In addition, we analyzed the production of reactive oxygen species (ROS) by the treatment of NL, NS, and NSP. All extracts reduced ROS production in a dose-dependent manner. And also, they increased heme oxygenase-1 (HO-1) protein expression and the nuclear translocation of nuclear respiratory factor 2 (Nrf2). In conclusion, our results suggest that Nelumbo nucifera has its anti-inflammatory activity via regulating MAPKs, NF-κB, and Nrf2/HO-1 pathways.

건조 상추 에탄올 추출물의 항염증 활성

이은주(Eun-Joo Lee),서유미(Yu-Mi Seo),김용현(Yong-Hyun Kim),정정욱(Chungwook Chung),성화정(Hwa-Jung Sung),손호용(Ho-Yong Sohn),박종이(Jong-Yi Park),김종식(Jong-Sik Kim) 한국생명과학회 2019 생명과학회지 Vol.29 No.3

상추는 가장 선호하는 녹색 채소 중 하나이다. 상추는 폴리페놀성 화합물을 비롯한 다양한 성분을 함유하고 있으며, 항균, 항산화, 항염증 등의 생리활성을 가지고 있는 것으로 알려져 있다. 본 연구에서는 건조상추의 에탄올 추출물(DLE)을 제조하고 이들의 항염증 활성을 연구하였다. DLE의 항염증 활성을 측정하기 위하여 LPS로 활성화된 마우스 대식세포 RAW 264.7 세포주에서 nitric oxide (NO) 생성을 측정하였다. DLE는 세포주의 생존에는 영향을 미치지 않으면서 NO 생산을 현저하게 저해하였다. DLE에 의해 염증 유전자인 iNOS와 COX-2의 유전자와 단백질의 발현이 모두 감소하였으며, 6개의 염증관련 cytokine 유전자(IL-1α, IL-1β, IL-1F6, TNF-α, CSF2, 그리고 CXCL10)의 발현이 모두 감소하였다. 또한, DLE의 처리는 MAPKs 경로의 인산화를 모두 저해하였으며, NF-κB p65의 핵으로의 이동을 저해하였다. 이러한 결과는 DLE의 항염증 활성은 MAPKs 경로와 NF-κB 경로를 조절함으로써 이루어짐을 시사한다. 또한, DLE는 농도의존적으로 reactive oxygen species (ROS)의 생산을 저해하였으며, hemeoxygenase-1 (HO-1) 단백질의 발현을 증가시켰으며, HO-1의 전사조절인자인 Nrf2의 핵으로의 이동을 증가시켰다. 결론적으로, 이러한 연구결과는 DLE가 염증관련 유전자의 발현을 감소시키며, MAPKs, NF-κB, 그리고 Nrf2/HO-1 등 다양한 경로를 조절함으로써 항염증 활성을 가지는 것을 제시한다. Lettuce (Lactuca sativa L.) is one of the most popular green leafy vegetables, and it contains various beneficial components including polyphenolic compounds and has been known to possess various biological functions such as anti-microbial, anti-oxidative, and anti-inflammatory activities. In the present study, we prepared ethanol extract of dried lettuce (DLE) and investigated its anti-inflammatory activity. To evaluate the anti-inflammatory activity of DLE, nitric oxide (NO) production was measured in LPS-activated mouse macrophage RAW 264.7 cells. DLE significantly suppressed NO production in these cells without affecting cell viabilities while resveratrol was used as a positive control. DLE dramatically decreased the expression of pro-inflammatory genes such as iNOS and COX-2 at the mRNA and protein levels and reduced the expression of several cytokines including IL-1α, IL-1β, IL-1F6, TNF-α, CSF2 and CXCL10. In addition, DLE suppressed phosphorylation of MAPKs and the nuclear translocation of NF-κB p65 indicating DLE shows its anti-inflammatory activity via regulating MAPKs pathway and NF-κB pathways. And also, DLE reduced the production of reactive oxygen species in a dose-dependent manner. DLE increased HO-1 protein expression, and also increased the nuclear translocation of Nrf2. Overall, our results suggest that lettuce down-regulate various pro-inflammatory genes and have its anti-inflammatory activity via regulating MAPKs, NF-κB, and Nrf2/HO-1 pathways.