Tobacco smoking induces cardiovascular mitochondrial oxidative stress, promotes endothelial dysfunction, and enhances hypertension

Abstract

Tobacco smoking is a major risk factor for cardiovascular disease and hypertension. It is associated with the oxidative stress and induces metabolic reprogramming, altering mitochondrial function. We hypothesized that cigarette smoke induces cardiovascular mitochondrial oxidative stress, which contributes to endothelial dysfunction and hypertension. To test this hypothesis, we studied whether the scavenging of mitochondrial H₂O₂ in transgenic mice expressing mitochondria-targeted catalase (mCAT) attenuates the development of cigarette smoke/angiotensin II-induced mitochondrial oxidative stress and hypertension compared with wild-type mice. Two weeks of exposure of wild-type mice with cigarette smoke increased systolic blood pressure by 17 mmHg, which was similar to the effect of a subpresssor dose of angiotensin II (0.2 mg·kg^-1·day^-1), leading to a moderate increase to the prehypertensive level. Cigarette smoke exposure and a low dose of angiotensin II cooperatively induced severe hypertension in wild-type mice, but the scavenging of mitochondrial H₂O₂ in mCAT mice completely prevented the development of hypertension. Cigarette smoke and angiotensin II cooperatively induced oxidation of cardiolipin (a specific biomarker of mitochondrial oxidative stress) in wild-type mice, which was abolished in mCAT mice. Cigarette smoke and angiotensin II impaired endothelium-dependent relaxation and induced superoxide overproduction, which was diminished in mCAT mice. To mimic the tobacco smoke exposure, we used cigarette smoke condensate, which induced mitochondrial superoxide overproduction and reduced endothelial nitric oxide (a hallmark of endothelial dysfunction in hypertension). Western blot experiments indicated that tobacco smoke and angiotensin II reduce the mitochondrial deacetylase sirtuin-3 level and cause hyperacetylation of a key mitochondrial antioxidant, SOD2, which promotes mitochondrial oxidative stress. NEW & NOTEWORTHY This work demonstrates tobacco smoking-induced mitochondrial oxidative stress, which contributes to endothelial dysfunction and development of hypertension. We suggest that the targeting of mitochondrial oxidative stress can be beneficial for treatment of pathological conditions associated with tobacco smoking, such as endothelial dysfunction, hypertension, and cardiovascular diseases.

Keywords: catalase; cigarette smoke; hypertension; mitochondria; oxidative stress; superoxide dismutase.

Fig. 1.

Cigarette smoke and angiotensin II (ANG II) cooperatively induce hypertension in C57BL/6 mice but not in mice expressing mitochondria-targeted catalase (mCAT). Mice had cigarette smoke nose-only exposure for 2 wk (4 cigarettes/day). Sham mice were housed in soft restraints for the duration of cigarette smoke treatment. Some mice received a subpressor low dose of ANG II (0.2 mg kg⁻¹⋅day⁻¹), which does not cause hypertension (61, 64). Data are expressed as means ± SD; n = 6. P values were calculated by two-way ANOVA. WT, wild type.

Fig. 2.

Cigarette smoke and angiotensin II (ANG II) cooperatively induce oxidation of mitochondrial cardiolipin in C57BL/6 mice but not in mice expressing mitochondria-targeted catalase (mCAT). Mice had cigarette smoke nose-only exposure (4 cigarettes/day), a subpressor low dose of ANG II infusion (0.2 mg⋅kg⁻¹⋅day⁻¹), or combined ANG II + smoke treatment. After 2 wk of treatment, mice were euthanized for liquid chromatography/mass spectrometry analysis of oxidized cardiolipin in the heart. Data are expressed as means ± SD; n = 6. §P < 0.01 vs. sham mice; #P < 0.05 vs. sham mice; **P < 0.01 vs. ANG II. L4CL, tetralinoleoylcardiolipin; oxL4CL, monohydroxy L4CL; WT, wild type.

Fig. 3.

Vasodilatation (A) and vascular superoxide (O₂^·−) production (B) in aortas isolated from mice treated with a combination of cigarette smoke and angiotensin II (ANG II; 0.2 mg⋅kg⁻¹⋅day⁻¹) for 2 wk. Vascular O₂^·− was measured by the 1-hydroxy-4-methoxy-2,2,6,6-tetramethylpiperidine hydrochloride spin probe and electron spin resonance, as we have previously described (11). Results are means ± SD; n = 6. *P < 0.01 vs. wild-type (WT) mice; **P < 0.05 vs. WT + ANG II/smoke. mCAT, mice expressing mitochondria-targeted catalase.

Fig. 4.

Effect of cigarette smoke and angiotensin II (ANG II) on acetylation of mitochondrial antioxidant SOD2 (SOD2-Ac), expression of SOD2, and sirtuin 3 (Sirt3). Mice had cigarette smoke exposure, ANG II infusion, or combined ANG II + smoke treatment. After 2 wk of treatment, mice were euthanized for kidney Western blot experiments (A). Cigarette smoke exposure and ANG II infusion induced SOD2 hyperacetylation (B), which diminished SOD2 activity (17). SOD2 expression was not significantly changed (C). SOD2 acetylation was accompanied by a moderate reduction of mitochondrial deacetylase sirtuin 3 (Sirt3; D). Data are normalized by β-actin level and expressed as means ± SE; n = 6. *P < 0.01 vs. sham mice; **P < 0.001 vs. ANG II.

Fig. 5.

Effect of cigarette smoke condensate (CSC) on mitochondrial superoxide (mito O₂^·−) production (A) and endothelial nitric oxide (NO; B) in the mouse aorta. Aortic sections isolated from sham wild-type mice were treated in DMEM tissue culture medium with CSC (40 µg/ml) or DMSO as vehicle (sham) for 24 h before electron spin resonance (ESR) measurements (insets) (10). *P < 0.01 vs. sham mice (n = 6). Fe(DETC)₂, ferrous-diethyldithiocarbamate.

Fig. 6.

Proposed role of mitochondrial oxidative stress in tobacco smoking-mediated endothelial dysfunction and hypertension. CNS, central nervous system; mCAT, mitochondria-targeted catalase; O₂^·−, superoxide; Sirt3, sirtuin 3.