Heme oxygenase-1 induction may explain the antioxidant profile of aspirin

doi:10.1016/S0006-291X(03)01504-3

Biochemical and Biophysical Research Communications

Volume 308, Issue 4, 5 September 2003, Pages 956-960

https://doi.org/10.1016/S0006-291X(03)01504-3 Get rights and content

Abstract

Aspirin is known to exert antioxidant effects by as yet unidentified mechanisms. In cultured endothelial cells derived from human umbilical vein, aspirin (30–300 μM) increased heme oxygenase-1 (HO-1) protein levels in a concentration-dependent fashion up to fivefold over basal levels. HO-1 induction was accompanied by a marked increase in catalytic activity of the enzyme as reflected by enhanced formation of both carbon monoxide and bilirubin. Pretreatment with aspirin or bilirubin at low micromolar concentrations protected endothelial cells from hydrogen peroxide-mediated toxicity. HO-1 induction and endothelial protection by aspirin were not mimicked by indomethacin, another inhibitor of cyclooxygenase. The nitric oxide (NO) synthase blocker L-NAME prevented aspirin-dependent HO-1 induction. These findings demonstrate that aspirin targets HO-1, presumably via NO-dependent pathways. Induction of HO-1 expression and activity may be a novel mechanism by which aspirin prevents cellular injury under inflammatory conditions and in cardiovascular disease.

Section snippets

Methods

Materials. Fetal bovine serum, cell culture media, and penicillin–streptomycin were obtained from Gibco (Eggenstein, FRG). Tin protoporphyrin-IX (SnPP) was purchased from Alexis Deutschland GmbH (Grünberg, FRG). The Chemiluminescence Western blotting kit and anti-rabbit peroxidase-conjugated secondary antibody were from Boehringer Mannheim (Mannheim, FRG). All other chemicals were bought from Sigma (Deisenhofen, FRG).

Cell culture. Human endothelial cells derived from umbilical cord were

Endothelial cell viability

A 12-h pretreatment with aspirin protected endothelial cells from hydrogen peroxide-mediated toxicity, whereas indomethacin failed to exert a cytoprotective action (Fig. 1). Endothelial protection by aspirin was mimicked by exogenous bilirubin which led to an almost complete reversal of hydrogen peroxide-induced toxicity (Fig. 2). In order to explore a potential involvement of HO products such as bilirubin in the observed protection by aspirin, the HO inhibitor SnPP was used. At a concentration

Discussion

The beneficial cardiovascular effects of aspirin are generally attributed to its immediate platelet inhibitory function. However, accumulating evidence suggests that aspirin may have additional biological properties on the vasculature that contributes to the reduction of ischemic cardiovascular events in patients with hypertension and atherosclerosis [27], [28]. These possible nonplatelet-mediated effects include the attenuation of atherosclerosis due to inhibition of vascular smooth muscle

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Pre-eclampsia (PE) is a pregnancy complication that can lead to maternal, fetal, and neonatal deaths in clinical practice. Accumulation of trophoblastic reactive oxygen species (ROS), which could result in oxidative stress and cell apoptosis, is considered to play an important role in PE pathology. It has been reported that aspirin has a positive effect on PE treatment in high-risk pregnant women.
In vitro, extravillous trophoblast cell line (HTR-8/SVneo) were treated with hydrogen peroxide (H₂O₂, 150 μM) after the presence of aspirin (90 and 120 μM) with or without GKT137831 (a Nox4 inhibitor, 20 μM). A series of experiments including CCK-8 assays, flow cytometry, biochemical testing, and Western Blotting etc. verified the protective effects and potential mechanisms of aspirin against oxidative stress-induced damage in PE.
Our results demonstrated that H₂O₂ induces oxidative stress and apoptosis in HTR8/SVneo cells. However, aspirin pretreatment rescue cell viability and reduce LDH activity of HTR-8/SVneo cells. Aspirin can suppress the ROS overproduction and MDA level while increase SOD content and CAT activity. In addition, aspirin pretreatment significantly alleviated cell apoptosis and suppressed the expression of Nox4 and its subunits (p22phox and p47phox) at protein and mRNA levels. The above results were more obvious after the combination of aspirin with GKT137831.
This study demonstrated that aspirin protects human trophoblasts against H₂O₂-induced oxidative stress and cell apoptosis via suppressing NADPH/ROS pathway. These findings provide novel insights for the application of aspirin as a protective and curative agent against PE.
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Citation Excerpt :
It inhibits the redox-sensitive transcription factor NF-κB, and the activity of COX1 and COX2, resulting in a strong reduction in the production of prostaglandins and TXA2 [232]. Moreover, aspirin reduces oxidative stress by inducing HO-1 activation [233]. Importantly, aspirin-acetylated COX2 may generate lipoxins from arachidonic acid [234], or “aspirin-triggered lipoxins” (ATLs or lipoxin-A4).
Preeclampsia (PE) is a multifactorial pregnancy disease, characterized by new-onset gestational hypertension with (or without) proteinuria or end-organ failure, exclusively observed in humans. It is a leading cause of maternal morbidity affecting 3–7% of pregnant women worldwide. PE pathophysiology could result from abnormal placentation due to a defective trophoblastic invasion and an impaired remodeling of uterine spiral arteries, leading to a poor adaptation of utero-placental circulation. This would be associated with hypoxia/reoxygenation phenomena, oxygen gradient fluctuations, altered antioxidant capacity, oxidative stress, and reduced nitric oxide (NO) bioavailability. This results in part from the reaction of NO with the radical anion superoxide (O₂^•−), which produces peroxynitrite ONOO^-, a powerful pro-oxidant and inflammatory agent. Another mechanism is the progressive inhibition of the placental endothelial nitric oxide synthase (eNOS) by oxidative stress, which results in eNOS uncoupling via several events such as a depletion of the eNOS substrate L-arginine due to increased arginase activity, an oxidation of the eNOS cofactor tetrahydrobiopterin (BH4), or eNOS post-translational modifications (for instance by S-glutathionylation). The uncoupling of eNOS triggers a switch of its activity from a NO-producing enzyme to a NADPH oxidase-like system generating O₂^•−, thereby potentiating ROS production and oxidative stress. Moreover, in PE placentas, eNOS could be post-translationally modified by lipid peroxidation-derived aldehydes such as 4-oxononenal (ONE) a highly bioreactive agent, able to inhibit eNOS activity and NO production. This review summarizes the dysfunction of placental eNOS evoked by oxidative stress and lipid peroxidation products, and the potential consequences on PE pathogenesis.
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For a better understanding of the effect of drugs and their interaction with cells and tissues, there is a need for in vitro and ex vivo model systems which enables studying these events. There are several in vitro methods available to evaluate the antioxidant activity; however, these methods do not factor in the complex in vivo physiology. Here we present an intestinal tissue modified oxygen electrode, used for the detection of the antioxidant effect of orally administered drugs in the presence of H₂O₂. Antioxidants are essential in the defense against oxidative stress, more specifically against reactive oxygen species such as H₂O_2. Due to the presence of native catalase in the intestine, with the tissue-based biosensor we were able to detect H₂O₂ in the range between 50 and 500 µM. The reproducibility of the sensor based on the calculated relative standard deviations was 15 ± 6%. We found that the O₂ production by catalase from H₂O₂ was reduced in the presence of a well-known antioxidant, quinol. This indirectly detected antioxidant activity was also observed in the case of orally administered drugs with a reported anti-inflammatory effect such as mesalazine and paracetamol, while no antioxidant activity was recorded with aspirin and metformin.
Translocator protein (18 kDa) (TSPO) ligands activate Nrf2 signaling and attenuate inflammatory responses and oxidative stress in human retinal pigment epithelial cells
2020, Biochemical and Biophysical Research Communications
Citation Excerpt :
Accordingly, we argue that this is likely the reason why XBD173 had less potent effects in inhibition of intracellular ROS levels in stressed human RPE cells compared to PK11195 and Ro5-4864. This possibility is consistent with the fact that, Heme oxygenase-1, the stress-response enzyme encoded by Hmox-1, seems to be essential for the therapeutic effects of a number of cytoprotective molecules including IL-10, aspirin and rapamycin; a phenomenon which has been referred to as the “HO-1 therapeutic funnel” [24–26]. The therapeutic effects of HO-1 expression are mediated by the degradation of heme to generate carbon monoxide (CO) and biliverdin, two molecules which possess potent anti-oxidant activities [27].
Degeneration of the retinal pigment epithelium (RPE) is a hallmark of atrophic age-related macular degeneration (AMD). Microglia mediated inflammatory responses and oxidative stress are critical pathophysiological processes in the onset and progression of RPE degeneration. Given the central role of the RPE, strategies to protect these cells from damage caused by oxidative stress and inflammation present a promising therapeutic approach to mitigate AMD. Ligands for the translocator protein (18 kDa) (TSPO) have been shown to confer protection against retinal inflammatory responses and neurodegeneration by acting primarily through retinal glia. However, despite RPE cells demonstrating strong TSPO expression, it remains unclear whether TSPO ligands could also inhibit inflammatory responses of RPE cells. Here, we investigated the influence of three different TSPO ligands XBD173, PK11195 and Ro5-4864 on inflammatory responses in human ARPE-19 cells triggered by supernatants from reactive human microglial cells and the lysosomal destabilizer, LLOMe. Our findings revealed that TSPO ligands significantly inhibited proinflammatory gene expression, inflammasome-mediated caspase-1 activation, lipid accumulation and intracellular ROS levels in stressed ARPE-19 cells. Notably, TSPO ligands induced activation of Nrf2 pathway and its downstream regulated genes in ARPE-19 cells, with Hmox-1 being the most strongly upregulated gene. Collectively, our study indicates that TSPO ligands can enhance the Nrf2 antioxidant pathway in RPE cells and protect them from cellular damage resulting from inflammation and oxidative stress.
Canagliflozin inhibits vascular smooth muscle cell proliferation and migration: Role of heme oxygenase-1
2020, Redox Biology
Recent cardiovascular outcome trials found that sodium-glucose cotransporter-2 (SGLT2) inhibitors reduce cardiovascular disease and mortality in type 2 diabetic patients; however, the underlying mechanisms are not fully known. Since the proliferation and migration of vascular smooth muscle cells (SMCs) contributes to the development of arterial lesions, we hypothesized that SGLT2 inhibitors may exert their beneficial cardiovascular effects by inhibiting the growth and movement of vascular SMCs. Treatment of rat or human aortic SMCs with clinically relevant concentrations of canagliflozin, but not empagliflozin or dapagliflozin, inhibited cell proliferation and migration. The inhibition of SMC growth by canagliflozin occurred in the absence of cell death, and was associated with the arrest of SMCs in the G₀/G₁ phase of the cell cycle and diminished DNA synthesis. Canagliflozin also resulted in the induction of heme oxygenase-1 (HO-1) expression, and a rise in HO activity in vascular SMCs, whereas, empagliflozin or dapagliflozin had no effect on HO activity. Canagliflozin also activated the HO-1 promoter and this was abrogated by mutating the antioxidant responsive element or by overexpressing dominant-negative NF-E2-related factor-2 (Nrf2). The induction of HO-1 by canagliflozin relied on reactive oxygen species (ROS) formation and was negated by antioxidants. Finally, silencing HO-1 expression partially rescued the proliferative and migratory response of canagliflozin-treated SMCs, and this was reversed by carbon monoxide and bilirubin. In conclusion, the present study identifies canagliflozin as a novel inhibitor of vascular SMC proliferation and migration. Moreover, it demonstrates that canagliflozin stimulates the expression of HO-1 in vascular SMCs via the ROS-Nrf2 pathway, and that the induction of HO-1 contributes to the cellular actions of canagliflozin. The ability of canagliflozin to exert these pleiotropic effects may contribute to the favorable clinical actions of the drug and suggest an extra potential benefit of canagliflozin relative to other SGLT2 inhibitors.

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^☆: Abbreviations: CO, carbon monoxide; HO, heme oxygenase; NO, nitric oxide; L-NAME, N^G-nitro-l-arginine-methyl-ester; SnPP, tin protoporphyrin-IX.

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Heme oxygenase-1 induction may explain the antioxidant profile of aspirin☆

Abstract

Section snippets

Methods

Endothelial cell viability

Discussion

Free Radic. Biol. Med.

FEBS Lett.

Biochem. Pharmacol.

Nitric Oxide

FEBS Lett.

J. Immunol. Methods

Anal. Biochem.

Thromb. Res.

Trends Pharmacol. Sci.

Aspirin as a therapeutic agent in cardiovascular disease

Special Writing Group, Circulation

Prostaglandins

Annu. Rev. Biochem.

Aspirin in cardiovascular disease

Drugs

The iron-binding and hydroxyl radical scavenging action of anti-inflammatory drugs

Xenobiotica

Aspirin protects low density lipoprotein from oxidative modification

Heart

Antioxidative properties of acetylsalicylic acid on vascular tissues from normotensive and spontaneously hypertensive rats

Circulation

Retrovirus-mediated HO gene transfer into endothelial cells protects against oxidant-induced injury

Am. J. Physiol.

Protective effects of transient HO-1 overexpression on susceptibility to oxygen toxicity in lung cells

Am. J. Physiol.

Heme oxygenase: protective gene or Trojan horse

Nat. Med.