Upregulation of heme oxygenase-1 as an adaptive mechanism for protection against crotonaldehyde in human umbilical vein endothelial cells

Crotonaldehyde, a highly reactive α, β-unsaturated aldehyde, is a ubiquitous environmental pollutant and a product of endogenous lipid peroxidation. It is also a major component of cigarette smoke and is present in many foods and beverages, and has also been linked to development of various diseases. Activation of endothelial cells by stimuli such as cigarette smoke is an important risk factor for cardiovascular diseases, including atherosclerosis. Heme oxygenase-1 (HO-1) is a protective antioxidant enzyme with a critical role in resistance to oxidative stress and other cellular functions. In this study, we examined the effects of crotonaldehyde on HO-1 induction and determined the signaling pathways in human umbilical vein endothelial cells (HUVECs). Inhibition of the protein kinase C-δ (PKC-δ) and p38 pathways resulted in significant blockage of crotonaldehyde-mediated HO-1 induction. Crotonaldehyde treatment caused a dramatic increase in translocation of NF-E2 related factor (Nrf2), leading to induction of HO-1. In addition, small interfering RNA knockdown of Nrf2 and treatment with the specific HO-1 inhibitor ZnPP exhibited an obvious increase of apoptosis of crotonaldehyde-treated HUVECs. Taken together, our results demonstrated that crotonaldehyde-induced HO-1 expression is mediated by the PKC-δ-p38 MAPK-Nrf2-HO-1 pathway in HUVECs, which is an adaptive response to oxidative stress.     

血红素加氧酶-1启动子区多态性与疾病易感性的关系

摘要血红加氧酶－1（HO－1）是一种诱导性抗氧化防御酶,可分解血红素释放出游离铁、一氧化碳和胆绿素,后者可迅速转化为胆红素,这三种代谢产物具有抗炎、抗氧化、抗增殖的作用。HO－1在大多数组织内呈低水平表达,可被多种伤害性刺激诱导产生高水平表达,从而在维持细胞内稳态方面起着重要作用。最近研究表明其基因启动子区5’端区域的2个突变可影响其转录水平,并且与多种疾病的发生发展密切相关,本文就此2个位点的多态性对HO－1的功能及其临床方面的意义作一综述。     

IL-37与过敏性疾病的研究进展

过敏性疾病是机体免疫系统对外来变应原产生的超敏反应性疾病, 目前糖皮质激素作为治疗过敏性疾病的一线用药, 其临床应用与治疗效果具有一定的局限性及不良反应。白细胞介素-37(interleukin-37, IL-37)作为天然的先天性和适应性免疫抑制剂, 具有双重抗炎功能, 可同时抑制炎症因子和激活抗炎因子的表达。研究表明, IL-37可通过多种作用机制及通路来维持过敏性疾病中免疫系统的动态平衡。因此, 本文将基于分析IL-37的本身特性, 对其在儿童常见的过敏性疾病中的作用机制作一综述, 旨在为进一步研究和临床诊疗提供依据。     

Heme oxygenase-1 in environmental toxin-induced lung disease

In the US, lung disease is the number three killer and accounts for one of every six deaths. Furthermore, more than 35 million US populations are now living with a chronic lung disease. Therefore, it is of urgent need to develop novel strategies that can protect against the development and progression of lung disease. Inhalation of air pollutants or environmental toxins induces inflammation and oxidative stress in the lung, resulting in tissue damage with subsequent decline in lung function. Heme oxygenase-1 (HO-1) is a stress response protein, which is highly inducible in response to pathological stimulation. Due to the cumulative effects of HO-1 on heme catabolism and the generation of biologically active downstream products, induction of HO-1 might serve as a protective mechanism against oxidative stress and inflammation-induced injury. Accumulating evidences have indicated a protective function of HO-1 against lung injury. This review highlights the roles of HO-1 in lung disease induced by environmental toxins such as cigarette smoke (CS), silica, and asbestos.     

The mechanism of heme oxygenase-1 action involved in the enhancement of neurotrophic factor expression

Heme oxygenase-1 (HO-1) is up-regulated in response to oxidative stress and catalyzes the degradation of pro-oxidant heme to carbon monoxide (CO), iron and bilirubin. Bilirubin is a potent antioxidant and neuroprotectant. Neurotrophic factors of BDNF and GDNF also play important roles in survival and morphological differentiation of dopaminergic neurons. We have previously found that HO-1 induction by adenovirus containing human HO-1 gene (Ad-HO-1) in substantia nigra of rat increases BDNF and GDNF expression. We here further examined the possible mechanism of HO-1 action involved in the enhancement of neurotrophic factor expression. Treatment of anti-BDNF/GDNF antibody significantly enhanced dopaminergic neuronal death, whereas Ad-HO-1 co-treatment was able to antagonize the apoptosis-inducing effect of these antibodies. The confocal imaging shows that HO-1 induction appeared in dopaminergic neuron, astrocyte and microglia at 24 h after injecting Ad-HO-1. HO-1 induced-BDNF/GDNF mRNA expression in substantia nigra was 26/21 folds of that of the contralateral Ad-injected side. The downstream product bilirubin increased GDNF expression through ERK and PI3K-Akt pathways, and also enhanced NFκB (p65 and p50) nuclear translocation in glia-enriched cultures. In addition, bilirubin also enhanced BDNF expression through similar pathway in cortical neuron-enriched cultures. We also examined the effect of another HO-1 product, CO, by using CO donor. [Ru(CO)₃Cl₂]₂ increased neurotrophic factor expression via sGC-PKG pathway in both neuron and glia. These results indicate that the downstream products of HO-1, i.e. bilirubin and CO, modulate BDNF and GDNF expression in neuron and astrocyte.     

Celastrol ameliorates HIV-1 Tat-induced inflammatory responses via NF-kappaB and AP-1 inhibition and heme oxygenase-1 induction in astrocytes

HIV-1 Tat causes extensive neuroinflammation that may progress to AIDS-related encephalitis and dementia. Celastrol possesses various biological activities such as anti-oxidant, anti-tumor, and anti-inflammatory activities. In this study, we investigated the modulatory effects of celastrol on HIV-1 Tat-induced inflammatory responses and the molecular mechanisms underlying its action in astrocytes. Pre-treatment of CRT-MG human astroglioma cells with celastrol significantly inhibited HIV-1 Tat-induced expression of ICAM-1/VCAM-1 and subsequent monocyte adhesiveness in CRT-MG cells. In addition, celastrol suppressed HIV-1 Tat-induced expression of pro-inflammatory chemokines, such as CXCL10, IL-8, and MCP-1. Celastrol decreased HIV-1 Tat-induced activation of JNK MAPK, AP-1, and NF-κB. Furthermore, celastrol induced mRNA and protein expression of HO-1 as well as Nrf2 activation. Blockage of HO-1 expression using siRNA reversed the inhibitory effect of celastrol on HIV-1 Tat-induced inflammatory responses. These results suggest that celastrol has regulatory effects on HIV-1 Tat-induced inflammatory responses by blocking the JNK MAPK-AP-1/NF-κB signaling pathways and inducing HO-1 expression in astrocytes.     

血红素加氧酶-1对慢性粒细胞白血病细胞凋亡效应的研究

目的探讨HO-1在慢性粒细胞白血病（以下简称慢粒）中的表达,研究其对抗细胞凋亡的效应,并研究抑制HO-1的表达对细胞凋亡的影响。方法探讨HO-1在慢性粒细胞白血病中的表达,研究其对抗细胞凋亡的效应,并研究抑制HO-1的表达对细胞凋亡的影响。结果RT-PCR结果显示骨髓血中HO-1的表达呈组成性,与对照组相比,诱导K562细胞HO-1的表达促进了细胞的生长（P＆lt;0.05）,并且这种表达成剂量依赖性,而抑制HO-1的表达促进了细胞的凋亡（P＆lt;0.05）,细胞的存活率只有对照组的30%。结论慢粒标本中HO-1的表达呈组成性,HO-1是一种可诱导型分子,并发挥抗凋亡效应,抑制HO-1的表达可能成为治疗慢粒的新方法。     

右美托咪啶对肾脏缺血再灌注损伤大鼠肾组织血红素氧合酶-1表达的影响

目的评价右美托咪啶对肾脏缺血再灌注损伤大鼠肾组织血红素氧合酶-1表达的影响。方法健康Wistar大鼠36只,雌雄不限,体重300~350g,随机分为3组:假手术组(S组)、肾脏缺血再灌注组(IR组)和右美托咪啶组(D组),各12只。采用动脉压夹夹闭双侧肾动脉60min、恢复灌注4h建立大鼠肾脏缺血再灌注模型。D组于夹闭双侧肾动脉前10min尾静脉注射右美托咪啶3μg/kg;IR组于夹闭双侧肾动脉前10min尾静脉注射等容量生理盐水;S组不夹闭双侧肾动脉,分离肾动脉后尾静脉注射等容量生理盐水。术后再灌注4h时处死大鼠取肾组织,采用PCR技术检测血红素氧合酶-1mRNA的表达,Westernblot法测定血红素氧合酶-1(HO-1)蛋白水平,光镜下观察肾组织病理学结果。结果与S组比较,IR组和D组肾组织血红素氧合酶-1mRNA和HO-1蛋白的表达上调(P<0.05);与IR组比较,D组肾组织血红素氧合酶-1mRNA和HO-1蛋白的表达上调(P<0.05),肾组织病理学损伤减轻。结论右美托咪啶减轻大鼠肾脏缺血再灌注损伤与其上调肾组织血红素氧合酶-1的表达有关。     

Regulation of heme oxygenase-1 expression and MAPK pathways in response to kaempferol and rhamnocitrin in PC12 cells

Oxidative stress has been considered as a major cause of cellular injuries in a variety of clinical abnormalities, especially neural diseases. Our aim of research is to investigate the protective effects and mechanisms of kaempferol and rhamnocitrin (kaempferol-7-methyl ether) on oxidative damage in rat pheochromocytoma PC12 cells induced by a limited supply of serum and hydrogen peroxide (H₂O₂). The current result demonstrated that kaempferol protected PC12 cells from serum deprivation-induced apoptosis. Pretreatment of cells with kaempferol also diminished intracellular generation of reactive oxygen species (ROS) in response to H₂O₂ and strongly elevated cell viability. RT-Q-PCR and Western blotting revealed that kaempferol and rhamnocitrin significantly induced heme oxygenase (HO)-1 gene expression. Addition of zinc protoporphyrin (Znpp), a HO-1 competitive inhibitor, significantly attenuated their protective effects in H₂O₂-treated cells, indicating the vital role of HO-1 in cell resistance to oxidative injury. While investigating the signaling pathways responsible for HO-1 induction, we observed that kaempferol induced sustained extracellular signal-regulated protein kinase 1/2 (ERK1/2) in PC12 cells grown in low serum medium; while rhamnocitrin only stimulated transient ERK cascade. Addition of U0126, a highly selective inhibitor of MEK1/2, which is upstream of ERK1/2, had no effect on kaempferol- or rhamnocitrin-induced HO-1 mRNA expression, indicating no direct cross-talk between these two pathways. Furthermore, both kaempferol and rhamnocitrin were able to persistently attenuate p38 phosphorylation. Taking together, the above findings suggest that kaempferol and rhamnocitrin can augment cellular antioxidant defense capacity, at least in part, through regulation of HO-1 expression and MAPK signal transduction.     

Compound C stimulates heme oxygenase-1 gene expression via the Nrf2-ARE pathway to preserve human endothelial cell survival

We recently identified adenosine monophosphate-activated protein kinase (AMPK) as a novel inducer of heme oxygenase-1 (HO-1) and surprisingly found that compound C (6-[4-(2-piperidin-1-yl-ethoxy)-phenyl]-3-pyridin-4-yl-pyrazolo[1,5-a] pyrimidine), a cell-permeable inhibitor of AMPK, could also elevate HO-1 suggesting other AMPK-independent actions for this agent. In this study, we investigated the biochemical mechanism by which compound C stimulates HO-1 expression in human endothelial cells (ECs) and determined the biological significance of the induction of HO-1 by compound C in these cells. Compound C stimulated a concentration- and time-dependent increase in HO-1 expression and an increase in HO-1 promoter activity that was abrogated by mutating the antioxidant responsive elements (AREs) in the HO-1 promoter or by overexpressing a dominant negative mutant of NF-E2-related factor 2 (Nrf2). Compound C also stimulated Nrf2 expression this was associated with an increase in the production of reactive oxygen species and with a decline in intracellular glutathione levels. Interestingly, the glutathione donor N-acetyl-l-cysteine or the NADPH oxidase inhibitor apocynin blocked the induction of HO-1 by compound C. Finally, compound C stimulated EC death and this was potentiated by silencing HO-1 expression and reversed by the administration of CO, biliverdin, or bilirubin. In conclusion, this study demonstrates that compound C stimulates HO-1 gene expression in human vascular endothelium via the activation of the Nrf2/ARE signaling pathway to counteract compound C-mediated cell death. The ability of compound C to induce HO-1 expression may contribute to the pleiotropic actions of this agent and suggest caution when using compound C to probe for AMPK functions.     

血红素氧合酶-1/一氧化碳系统在肺纤维化大鼠中的表达及药物干预

目的研究血红素氧合酶-1（HO-1）/一氧化碳（CO）系统在肺纤维化大鼠中的表达及药物干预影响,为肺纤维化的诊治提供一种新方法。方法应用健康Wistar大鼠60只,随机分为4组,分别为对照组、肺纤维化组（纤维化组）、肺纤维化加HO-1特异激动剂组（加强组）、肺维化加HO-1特异抑制剂组（抑制组）。检测各个模型组中大鼠肺组织与血清中HO-1/CO体系表达情况,分析HO-1/CO体系在大鼠肺纤维化中的作用,以及药物对肺纤维化程度的影响。结果在对照组中,HO-1与COHb在大鼠肺组织和血清中低表达,且两者差异无统计学意义（P〉0.05）,在单纯组、加强组、抑制组中,与对照组相比,HO-1与COHb在大鼠肺组织和血清中均高表达,以加强组为重,抑制组最低,两者差异具有统计学意义（P〈0.05）。结论HO-1/CO系统参与了大鼠肺纤维化形成过程,通过对其的干预,可以改变大鼠肺纤维化的程度,为肺纤维化的诊断与治疗的提供新途径。     

Tetrachlorobenzoquinone induces acute liver injury,up-regulates HO-1 and NQO1 expression in mice model: The protective role of chlorogenic acid

Tetrachlorobenzoquinone (TCBQ) is an active metabolite of pentachlorophenol (PCP). Although PCP has been investigated extensively, there are only a few reports describing the toxicity effect of TCBQ, and no report regarding TCBQ-induced liver injury in vivo. In the current study, we aimed to examine the acute hepatic toxicity of TCBQ in the mice model. Chlorogenic acid (CGA) exhibits promising antioxidant activity in the past studies, thus, the second aim of this study was to evaluate the protective effect of CGA on TCBQ-induced liver injury. Our results indicated TCBQ-intoxication caused marked liver cell necrosis and inflammation but not apoptosis, and this damage was alleviated by CGA treatment. Meantime, TCBQ-intoxication enhanced serum ALT, AST activities, TBIL content, hepatic oxidative stress and lipid peroxidation, decreased GSH content and inhibited the activities of antioxidant enzymes. Western blot and immunohistochemical analysis showed that TCBQ marked up-regulated HO-1 and NQO1 expression. On the other hand, pretreatment of CGA reduced TCBQ-induced liver damage remarkably. Taking together, these results revealed that TCBQ has strong hepatic toxic effect, and at least a part of this effect is initiated by free radical and relieved with CGA administration.     

Dipyridamole inhibits lipopolysaccharide-induced cyclooxygenase-2 and monocyte chemoattractant protein-1 via heme oxygenase-1-mediated reactive oxygen species reduction in rat mesangial cells

Dipyridamole contributes to its beneficial effects on inflammatory responses in many cell types. The anti-inflammatory mechanisms of dipyridamole on glomerular mesangial cells are mostly uncharacterized. In this study, we monitored the influence of dipyridamole on the expression levels of cyclooxygenase-2 (COX-2) and monocyte chemoattractant protein-1 (MCP-1) in rat mesangial cells stimulated with lipopolysaccharide. Dipyridamole was found to inhibit lipopolysaccharide-induced COX-2 and MCP-1 expression, and reduced lipopolysaccharide-induced reactive oxygen species generation in rat mesangial cells. This inhibitory effect of dipyridamole is independent on cyclic AMP and cyclic GMP increase. Tin protoporphyrin IX (SnPP), a heme oxygenase-1(HO-1) inhibitor, blocked the inhibitory effect of dipyridamole on lipopolysaccharide-induced COX-2 and MCP-1 expression. By applying specific inhibitors in rat mesangial cells, ERK1/2 and p38 MAPK signaling pathways were demonstrated to be involved in the lipopolysaccharide-induced inflammatory responses, and were inhibited by SnPP and N-acetylcysteine treatment. Additionally, dipyridamole was also found to upregulate HO-1 in rat mesangial cells. Therefore, our data suggest that dipyridamole inhibits the expression of COX-2 and MCP-1 in lipopolysaccharide-treated rat mesangial cells via HO-1-mediated reactive oxygen species reduction.     

Role of heme oxygenase-1 in vascular disease

Great attention has been placed on the protective role of heme oxygenase-1 (HO-1) for several vascular diseases such as atherosclerosis. HO-1, by exerting anti-inflammatory, antiproliferative, anti-apoptotic and anti-oxidant effects on the vasculature, protects against atherosclerosis. The precise underlying mechanisms for HO-1-based protection are not yet completely understood, but appear to involve the protective effects of HO-1 by-products, carbon monoxide (CO), biliverdin/bilirubin and free iron. Among the HO-1 by-products, CO has been shown to mimic some protective actions of HO-1, specifically, in vascular system. There is evidence supporting that HO-1-derived CO also interacts with other gaseous molecules, such as nitric oxide (NO) and hydrogen sulfide (H2S) that may relate to either vascular protection or injury. CO, NO and H2S not only exert comparable biological actions but also compete with and are antagonists with each other for maintaining vascular homeostasis. This review will highlight the protective roles of HO-1/CO in vascular injury/disease, and emphasize the potential roles of CO in possible interplay among three gaseous molecules, which may be important to explore the overall protective roles of HO-1/CO system in the pathogenesis of human vascular disease.     

Flavonoids from silkworm droppings and their promotional activities on heme oxygenase-1

A new flavane glucoside, 7,2′-dihydroxy-8-hydroxyethyl-4′-methoxyflavane-2′-O-β-d-glucopyranoside (3), along with three known flavonoids, 7,2′-dihydroxy-8-prenyl-4′-methoxyflavane (1), euchrenone a₇ (2), and 7,2′-dihydroxy-8-prenyl-4′-methoxy-2′-O-β-d-glucopyranosylflavane (4), was isolated from silkworm droppings. The structures of the compounds were elucidated on the basis of 1D and 2D NMR spectroscopic analyses and optical rotational characteristics. The compounds isolated from silkworm droppings were evaluated for their effects on heme oxygenase-1 (HO-1) activity. Compounds 1 and 3 increased the expression of HO-1 in HepG2 cells. HO-1 is an antioxidant enzyme that catabolizes heme to carbon monoxide, free iron, and biliverdin, all of which are involved in the suppression of inflammatory mediators.     

Induction of heme oxygenase-1 with hemin attenuates hippocampal injury in rats after acute carbon monoxide poisoning

Carbon monoxide (CO) poisoning is a major cause of brain injury and mortality; delayed neurological syndrome (DNS) is encountered in survivors of acute CO exposure. The toxic effects of CO have been attributed to oxidative stress induced by hypoxia. Heme oxygenase-1 (HO-1) is the inducible heme oxygenase isoform, and its induction acts as an important cellular defense mechanism against oxidative stress, cellular injury and disease. In this study, we examined the functional roles of HO-1 induction in a rat model of CO-exposured hippocampal injury. We report that acute CO exposure produces severe hippocampal injury in rats. However, hemin pretreatment reduced both the CO-induced rise in hippocampal water content and levels of neuronal damage in the hippocampus; survival rates at 24 h were significantly improved. Upregulation of HO-1 by hemin pretreatment resulted in a significant decrease in hippocampal levels of malondialdehyde (MDA), a marker of oxidative stress; levels of pro-apoptotic caspase-3 were also reduced. In contrast, inhibition of HO activity by administration of tin protoporphyrin IX (SnPP, a specific inhibitor of HO) abolished the neuroprotective effects of HO-1 induction. These data suggested that the upregulation of endogenous HO-1 expression therefore plays a pivotal protective role in CO neurotoxicity. Though the precise mechanisms underlying hemin-mediated HO-1 induction and neuroprotection are not known, these may involve the anti-oxidant and anti-apoptotic effects of HO-1 enzyme activity.     

Expression of heme oxygenase-1 and regulation by cytokines in human osteoarthritic chondrocytes

Heme oxygenase-1 (HO-1) is implicated in the protection against tissue injury. We investigated the expression of this protein in cartilage sections and chondrocytes obtained from osteoarthritic patients. HO-1 was immunodetected in preparations from cartilage and also in chondrocytes cultured in the absence of stimulation. We found that HO-1 can be modulated by cytokines since the pro-inflammatory cytokines interleukin (IL)-1β, IL-17 and tumour necrosis factor-α (TNF-α) down-regulated this protein, whereas the anti-inflammatory cytokine IL-10 exerted the opposite effect. Our results suggest a role for HO-1 as part of protective mechanisms against tissue injury in human cartilage.     

MicroRNA changes associated with atypical CYP1A1 inducer BMS-764459

The corticotrophin releasing factor (CRF) receptor I antagonist, BMS-764459 (evaluated as a potential treatment of affective disorders), was orally dosed to female Sprague-Dawley rats once daily for 2 weeks (vehicle control or 175 mg/kg/day). To investigate the mechanism of BMS-764459-related liver weight increases, total liver RNA was isolated and evaluated for mRNA gene expression by microarray analysis (assessing the expression of approximately 24,000 genes) from snap-frozen tissue. Subsequently, mRNA and miRNA (microRNA) were also analyzed 5 years later from FFPE (Formalin Fixed Paraffin Embedded) samples via RT-PCR (about 800 miRNA evaluated). Genomic analyses showed that BMS-764459 induces AhR target genes with additional inductions of CYP2B, CYP3A, and Abcc3 consistent with the gene expression pattern of atypical CYP1A1 inducers. Analysis of miRNA expression identified a number of significantly affected miRNAs. To further evaluate their role in atypical CYP1A1 induction, an in silico evaluation of differentially expressed miRNA was performed and their putative mRNA 3′-UTR (untranslated region) binding sequences were evaluated. MiR-680 and miR-29a were identified as potential regulators and biomarkers of atypical CYP1A1 induction by regulating Abcc3, CYP3A and CYP2B as well as a number of AhR targeted genes.