Nicotine and the renin-angiotensin system
Joshua M. Oakes
1Department of Physiology, Louisiana State University Health Sciences Center, New Orleans, Louisiana
Robert M. Fuchs
2Department of Pharmacology and Experimental Therapeutics, Louisiana State University Health Sciences Center, New Orleans, Louisiana
Jason D. Gardner
1Department of Physiology, Louisiana State University Health Sciences Center, New Orleans, Louisiana
Eric Lazartigues
2Department of Pharmacology and Experimental Therapeutics, Louisiana State University Health Sciences Center, New Orleans, Louisiana
Xinping Yue
1Department of Physiology, Louisiana State University Health Sciences Center, New Orleans, Louisiana
Corresponding author.Abstract
Cigarette smoking is the single most important risk factor for the development of cardiovascular and pulmonary diseases (CVPD). Although cigarette smoking has been in constant decline since the 1950s, the introduction of e-cigarettes or electronic nicotine delivery systems 10 yr ago has attracted former smokers as well as a new generation of consumers. Nicotine is a highly addictive substance, and it is currently unclear whether e-cigarettes are “safer” than regular cigarettes or whether they have the potential to reverse the health benefits, notably on the cardiopulmonary system, acquired with the decline of tobacco smoking. Of great concern, nicotine inhalation devices are becoming popular among young adults and youths, emphasizing the need for awareness and further study of the potential cardiopulmonary risks of nicotine and associated products. This review focuses on the interaction between nicotine and the renin-angiotensin system (RAS), one of the most important regulatory systems on autonomic, cardiovascular, and pulmonary functions in both health and disease. The literature presented in this review strongly suggests that nicotine alters the homeostasis of the RAS by upregulating the detrimental angiotensin-converting enzyme (ACE)/angiotensin (ANG)-II/ANG II type 1 receptor axis and downregulating the compensatory ACE2/ANG-(1–7)/Mas receptor axis, contributing to the development of CVPD.
INTRODUCTION
Cigarette smoking is the single most important risk factor for the development of cardiovascular and pulmonary diseases (CVPD), and smokers are two to four times more likely to develop CVPD than nonsmokers (132a, 73). As summarized in The Health Consequences of Smoking-50 Years of Progress: A Report of the Surgeon General, the evidence is sufficient to infer a causal relationship between smoking and coronary heart disease, atherosclerotic aortic aneurysm, cerebrovascular disease/stroke, acute respiratory illnesses, chronic obstructive pulmonary disease (COPD), exacerbation of asthma, lung cancer, and all-cause mortality (132a).
Although cigarette smoking has been in constant decline since the 1950s, the introduction of electronic-cigarettes (e-cig) or electronic nicotine delivery systems 10 yr ago has attracted former smokers as well as a new generation of consumers. Nicotine is a highly addictive substance, and it is currently unclear whether e-cig are “safer” than regular cigarettes or whether they have the potential to reverse the health benefits, notably on the cardiopulmonary system, acquired with the decline of tobacco smoking. Of great concern, nicotine inhalation devices are becoming popular among young adults and youths (135), emphasizing the need for awareness and further study of the potential cardiopulmonary risks of nicotine and associated products. The current review highlights the interaction between nicotine and the renin-angiotensin system (RAS), one of the most important regulatory systems on autonomic, cardiovascular, and pulmonary functions in both health and disease.
OVERVIEW OF THE RAS SYSTEM
RAS dysfunction has been implicated in CVPD, including arterial and pulmonary hypertension, congestive heart failure, endothelial dysfunction, myocardial infarction, acute and chronic lung diseases, atherosclerosis, organ fibrosis, and cardiorenal metabolic syndrome (63, 76, 85, 88, 104, 124). The RAS is a peptide hormone system composed of several enzymes and inactive and active peptides, which altogether play an important role in neurovascular and cardiopulmonary physiology (47, 88). In addition to the well-known systemic RAS, local systems contributing to specific organ functions have been described for every major organ and tissue (3, 15, 37, 40, 42, 64, 66, 87, 110, 145). In the classic representation of the RAS (Fig. 1), angiotensinogen is hydrolyzed by renin to produce angiotensin (ANG) I, which is then converted by angiotensin-converting enzyme (ACE) into the biologically active ANG II. More recently, renin- and ACE-independent formation of ANG II have also been reported (109, 131, 140). The ANG II type 1 receptor (AT1R) is the primary receptor that mediates most of the effects of ANG II, including vasoconstriction, water, and salt reabsorption and enhanced sympathetic drive. The ANG II type 2 receptor (AT2R) has also been identified in various tissues, and its activation opposes AT1R-mediated effects and therefore is thought to be involved in the compensatory responses, notably via nitric oxide release and antigrowth properties (14, 17, 25, 35, 49, 51–53, 67). The discovery of ACE2 in 2000 has paved the way for the recognition of a new compensatory arm of the RAS (34, 130). ACE2 is a monocarboxypeptidase, impervious to ACE inhibitors. ACE2 cleaves ANG I to generate ANG-(1–9), which is then converted to the vasodilatory peptide ANG-(1–7) by ACE or other peptidases. With higher efficiency, ACE2 also directly metabolizes ANG II to form ANG-(1–7). This heptapeptide has opposite properties to those of ANG II, promoting vasodilation and exerting antiproliferative and antihypertrophic effects by acting through the Mas receptor (MasR) (116). Some reports have described ANG-(1–7) activation of AT2R as well (136). By cleaving ANG II into ANG-(1–7), ACE2 plays a pivotal role in the compensatory ACE2/ANG-(1–7)/MasR axis of the RAS by counterbalancing the deleterious actions of the ACE/ANG II/AT1R arm (21, 45, 55, 71, 117, 118, 145). In addition to the above components of the RAS, ANG II is further metabolized to ANG III by aminopeptidase A, which is then converted to ANG IV by aminopeptidase N (50). ANG II can also be converted to angiotensin A by mononuclear leukocyte-derived aspartate decarboxylase, leading to the formation of alamandine, which has been shown to bind to the Mas-related G protein-coupled receptor D (68).
Overview of the renin-angiotensin system (RAS) and the interaction between nicotine and the RAS. ACE, angiotensin-converting enzyme; APA, aminopeptidase A; APN, aminopeptidase N; AT1R, angiotensin II type 1 receptor; AT2R, angiotensin II type 2 receptor; MasR, Mas receptor; MLDAD, mononuclear leukocyte-derived aspartate decarboxylase.
NICOTINE AND THE RAS IN CARDIOVASCULAR DYSFUNCTION
Nicotine and the RAS in blood pressure regulation.
The interaction between nicotine exposure and components of the RAS has long been recognized. It was shown 30 yr ago that cigarette smoking or nicotine inhalation in human volunteers led to acute increases in both systolic and diastolic blood pressure (BP) (16, 54, 56) and these effects were accompanied by increased plasma ACE activity (54), indicating the involvement of the RAS. Activation of the RAS by chronic cigarette smoking was supported by a well-controlled study in human monozygotic twins, which showed the presence of higher plasma renin activity and elevated plasma aldosterone levels in the smoking twin (with at least 10 yr of uninterrupted cigarette use) compared with the nonsmoking twin, both at rest and during exercise (86). In the Ludwigshafen Risk and Cardiovascular Health (LURIC) Study, plasma renin and aldosterone levels were not found to be significantly different between never smokers and active smokers (32). This study, however, was confounded by differences in age, gender, comorbidities and the use of antihypertensive drugs between the two groups, as well as the lack of exact smoking history in the active smokers (32). Studies in laboratory animals support the activation of RAS by cigarette smoke or nicotine. Chronic nicotine administration through osmotic minipumps has been shown to elevate plasma renin activity in rats subjected to a high-salt diet (120), and cigarette smoke and/or nicotine administration increase plasma ACE activity with increased conversion of ANG I to ANG II (7, 80, 126).
Increased ACE activity also contributes to impaired vascular relaxation observed in smokers. Irbesartan, an AT1R antagonist (also known as angiotensin receptor blocker), was found to reduce arterial stiffness in hypertensive patients (134). When stratified by smoking status, smokers were found to have stiffer arteries before treatment, and irbesartan was able to reduce arterial stiffness to a greater extent in smokers than in nonsmokers, indicating that over-activity of the RAS contributes to increased arterial stiffness in smokers (134). With the use of the hand vein technique, smokers were found to have impaired venodilation in response to bradykinin compared with nonsmokers (18). Infusion of enalaprilat, an ACE inhibitor, restored bradykinin-induced relaxation in smokers to values found in nonsmokers (18). This observation suggests that increased vascular metabolism of bradykinin (another function of ACE) takes place in veins of smokers and that vascular RAS may play a key role in smoking-induced endothelial dysfunction.
Studies of the effects of e-cig on BP are limited. One study examined BP in smokers with arterial hypertension who switched to e-cig and found that patients who switched from tobacco smoking to e-cig had reduced resting BP and better BP control (113). This study suggests that e-cig may be less harmful than conventional cigarettes in hypertensive smokers but does not prove that e-cig has no adverse health consequences. Indeed, arterial BP was increased acutely after both conventional cigarette and e-cig use in habitual smokers, even though the increase was less with e-cig (146). Importantly, in young healthy nonsmokers, acute inhalation of e-cig vapor increases both systolic and diastolic BP and heart rate (24). In another study, increased heart rate variability was reported following exposure to e-cig with nicotine, but not to e-cig without nicotine, indicating a shift in cardiac sympatho-vagal balance toward sympathetic predominance (101). In young, normotensive nonsmokers, Fogt et al. (48) found that e-cig inhalation preferentially increased both resting and exercise diastolic BP 40 min following nicotine inhalation. A recent prospective 3.5-yr observational study by Polosa et al. (112) found that in a population of healthy e-cig users who had never smoked conventional cigarettes, there was no change in systolic or diastolic BP compared with healthy never smokers and non-e-cig users. The authors thus concluded that daily e-cig use is not associated with adverse health concerns. However, the small sample size (6 daily e-cig users consuming nicotine-containing e-liquid), relative young age (26.6 ± 6.0 yr), and short duration of e-cig use (average of 4.2 yr in total) make this conclusion premature.
Maternal cigarette smoking is associated with an increased risk of cardiovascular diseases in postnatal life. Perinatal nicotine exposure has been shown to promote RAS dysfunction in the adult offspring (143). In this study, nicotine was administered to pregnant rats via subcutaneous osmotic minipumps from gestational day 4 to postnatal day 10, and BP and vascular responses to ANG II were measured in 5-mo-old adult offspring. Perinatal nicotine had no effect on baseline BP but significantly increased ANG II-stimulated BP increase in male but not in female offspring. In male adult offspring, perinatal nicotine significantly increased arterial media thickness and potentiated ANG II-induced contractions of aorta and mesenteric artery, which were blocked by losartan, a selective AT1R antagonist. In addition, perinatal nicotine exposure increased AT1R but decreased AT2R protein expression in the aorta of adult male offspring, resulting in an imbalance of the AT1R/AT2R ratio in favor of the vasoconstrictor component. Epigenetic regulation of gene expression has been proposed to be the mechanism by which nicotine alters the expression of ANG receptors. Quantitative methylation-specific PCR using aortic rings isolated from male adult offspring showed decreased methylation in the promoter regions of AT1R but increased methylation in the promoter region of AT2R (141), consistent with increased AT1R and decreased AT2R mRNA expression (141, 143). Estrogen plays a key role in the sex difference of perinatal nicotine-induced programming of vascular dysfunction. In ovariectomized female progeny, perinatal nicotine exposure enhanced ANG II-induced BP response and vascular contraction similarly as in male offspring, and these heightened responses were abrogated by 17β-estradiol replacement (142).
Nicotine and the RAS in cardiac remodeling.
In the heart, AT1R activation by ANG II leads to cardiac fibrosis, cardiomyocyte hypertrophy, and inflammation (38), whereas AT2R acts as a counterbalance to AT1R, performing antifibrotic, antihypertrophic, and anti-inflammatory roles (89, 114). The heart is composed of many different cell types, including cardiomyocytes, fibroblasts, endothelial cells, and smooth muscle cells, each of them exhibiting a different RAS expression profile. ACE is expressed in vascular endothelial cells, fibroblasts, and cardiomyocytes (albeit at low levels), whereas ACE2 is expressed primarily in vascular endothelial cells (31, 46, 58, 128). All cardiac cell types express AT1R and AT2R (8, 132, 141); however, the expression of AT2R is low in the healthy heart, and the primary effects of ANG II are mediated through interactions with AT1R under normal conditions. The MasR is also expressed in the heart and localized to the vasculature, cardiomyocytes, and cardiac fibroblasts (72).
Nicotinic acetylcholine receptors are widely expressed in the heart (36, 92), and there is evidence that chronic cigarette smoking or nicotine exposure exacerbates pathological changes in the heart secondary to hypertension or other cardiac stressors. Cigarette smoke was found to accelerate the progression of cardiac hypertrophy and heart failure in spontaneously hypertensive rats (99). Using the chronic volume overload (VO) model in rats, we have shown that cigarette smoke significantly increased VO-induced left ventricular dilatation, prevented compensatory wall thickening, and depressed fractional shortening (11). At the molecular level, cigarette smoke blunted compensatory collagen expression, exacerbated VO-induced matrix metalloproteinase (MMP)-9 and tissue inhibitor of metalloproteinase 1 expression, and blocked the compensatory increases in hypoxia inducible factor-1α, vascular endothelial growth factor, and transforming growth factor-β in the VO-stressed heart (11). A recent study by Vang et al. (133) showed that 6-wk cigarette smoke exposure in mice specifically induced right ventricular dysfunction and fibrosis in the absence of right ventricular hypertrophy and pulmonary vascular remodeling. Both cigarette smoke extract and nicotine increased cardiac fibroblast proliferation and collagen synthesis in rat fibroblasts. These effects were mediated through α7-nicotinic acetylcholine receptors and were dependent on protein kinase C (PKC)-α/δ and reduced p38-mitogen-activated protein kinase (MAPK) phosphorylation (133). Possible cardiotoxic interaction between nicotine and the RAS was also demonstrated in a mouse model of systemic hypertension (22). In this study, nicotine and ANG II were delivered through subcutaneous osmotic minipumps either individually or in combination. Nicotine was shown to exacerbate cardiovascular remodeling induced by ANG II, which included increased heart rate, increased myocardial MMP-2 expression, and increased thickening of the aortic wall (22).
The long-term effects of e-cig on cardiac function are currently unknown. A recent study by Farsalinos et al. (41) found that acute e-cig inhalation did not alter left ventricular function in healthy e-cig users, as compared with healthy heavy smokers who exhibited a delay in myocardial relaxation following acute smoking.
Nicotine and the RAS in other vascular events.
Both nicotine and RAS have been implicated in the pathogenesis of vascular calcification and abdominal aortic aneurysms (AAA). Sui et al. (127) found that ANG-(1–7), delivered through osmotic minipumps, prevented vascular calcification induced by vitamin D3 plus nicotine in rats. Activation of the RAS was observed in calcified aortas, and the administration of ANG-(1–7) resulted in reduced ACE and AT1R mRNA expression in the aorta, reduced plasma ACE activity, reduced osteogenic transition of vascular smooth muscle cells (VSMCs), and reduced calcium deposition in the aortas (127). RAS activation is also involved in the development of AAA, and smoking is an important risk factor associated with the development, expansion, and rupture of AAA (29, 95). Acute infusion of nicotine markedly increased the incidence of AAA in apolipoprotein E knockout mice, and this effect was likely mediated by nicotine-induced upregulation of MMP-2 in VSMCs and the activation of AMP-activated protein kinase-α2 (139). Similarly, Guo et al. (57) found that mice coadministered with nicotine and ANG II had increased incidence of AAA accompanied by increased expression of MMP-2/9 and proinflammatory chemokines.
NICOTINE AND THE RAS IN THE NERVOUS SYSTEM
Nicotine classically acts in the brain by binding to neuronal nicotinic acetylcholine receptors, which are virtually ubiquitous throughout the central nervous system and play an important excitatory role in autonomic ganglia (27). Inhaled nicotine has rapid access to neural tissue (9), and in vivo studies have shown that chronic nicotine exposure increases blood-brain barrier permeability (62). Within the brain, nicotinic receptors have been extensively studied in the context of the mesolimbic system, where nicotine modulates release of glutamate and other neurotransmitters to promote behavioral dependence (26, 27, 83). These receptors also influence activity of autonomic regions such as the hypothalamic paraventricular nucleus (PVN) and promote a sympathomimetic stress response through the hypothalamic-pituitary-adrenal axis (2).
Virtually every component of the systemic RAS exists in the brain (77, 145). As the blood-brain barrier prevents systemic ANG II from readily accessing neural tissue, the brain RAS functions independently of the systemic RAS (119). Endogenous ANG II has been detected in the aforementioned PVN of the hypothalamus as well as in the nucleus of the solitary tract in the brainstem (93), the latter of which plays a major role in the regulation of baroreceptor reflex function (100). Stereotaxic injection of ANG II into the PVN of rodents has been shown to acutely increase BP, and administration of losartan blocked this effect, highlighting the importance of the brain RAS in PVN-mediated autonomic control (4). Moreover, PVN-specific overexpression of ACE2 in mice has been shown to reduce AT1R levels and attenuate the acute prohypertensive effects of exogenous ANG II administration (125).
A specific mechanism by which nicotine could influence autonomic nuclei is through interaction with the brain RAS (145). Adverse addictive and cardiovascular sequelae associated with chronic smoking are mediated, at least in part, by nicotine-induced activation of nicotinic acetylcholine receptors within the striatal dopaminergic (39) and hypothalamic noradrenergic system (61, 122), respectively. Narayanaswami et al. (102) examined the role of AT1R and AT2R in nicotine-evoked dopamine and norepinephrine release from striatal and hypothalamic slices and showed that losartan concentration dependently inhibited nicotine-evoked dopamine and norepinephrine release, whereas PD123319 (AT2R antagonist) did not alter nicotine-evoked norepinephrine release from hypothalamus but potentiated nicotine-evoked dopamine release from the striatal tissue. The above results indicate that AT1R is involved in nicotine-evoked release of both dopamine and norepinephrine, and AT1R activation in striatum is counterbalanced by AT2R. Furthermore, nicotine exposure alters the expression and activity of the brain RAS. In primary cultures of neurons and glial cells isolated from brainstem and hypothalamus of 1-day-old rats, nicotine treatment resulted in increased expression of AT1R but decreased expression of ACE2, and these nicotine effects were greater in cells isolated from spontaneously hypertensive rats compared with Wistar-Kyoto rats (43, 44). The above findings provide strong evidence that interaction between nicotine and the brain RAS may contribute to the sympatho-mimetic actions of nicotine as well as AT1R-mediated neurogenic hypertension (144).
With regard to the catecholamines, nicotine has been shown to induce the synthesis and release of norepinephrine/epinephrine from the adrenal medulla, an extension of the sympathetic nervous system. It was shown over 20 yr ago that nicotine modulates adrenal expression of tyrosine hydroxylase (the rate-limiting enzyme in the catecholamine biosynthetic pathway), dopamine β-hydroxylase (which converts dopamine to norepinephrine) and neuropeptide Y (which is coreleased with the catecholamines) (65). In vitro, both ANG II and nicotine promote catecholamine release from chromaffin cells (121), and pretreatment of adrenal tissue with nicotine potentiates catecholamine release in response to ANG II infusion (84).
Interaction between nicotine and the brain RAS is not limited to the hypothalamus, and another region of interest is the subfornical organ. Intracerebroventricular injection of ANG II and nicotine have both been shown to elevate expression of c-Fos (a marker of neuronal activation) in the subfornical organ and acutely promote water drinking behavior (106, 115). Patch-clamp recording of cells dissociated from this region revealed a subpopulation of neurons that have an excitatory response to both ANG II and nicotine (107). Future studies could clarify whether selective excitation of these neurons is associated with a sympathetic response such as elevated systemic BP and how nicotine and the RAS may modulate this response. Investigations of nicotine and the RAS in other autonomic brain regions would also be informative, especially in the baroreflex-regulating areas such as the rostral ventrolateral medulla and the nucleus of the solitary tract (19, 20).
Perinatal nicotine exposure alters the brain RAS and results in increased sympathetic activity in the adult offspring. In rats with prior in utero nicotine exposure, a reduction of baroreflex sensitivity in response to ANG II infusion was observed (143, 147). In addition, prenatal nicotine exposure increases blood norepinephrine levels at baseline as well as following intravenous administration of ANG II (147). Increased AT1R and decreased AT2R expression were observed in whole brain extracts from fetal and adult offspring rats that had been subjected to perinatal nicotine exposure (96). The downregulation of AT2R in the brain was shown to occur through nicotine-mediated hypermethylation of the AT2R promoter (90, 91).
NICOTINE AND THE RAS IN THE LUNG
The lung is the first organ to encounter the inhaled nicotine. Many cell types in the lung express nicotinic acetylcholine receptors, including bronchial epithelial cells, type II alveolar epithelial cells, alveolar macrophages, pulmonary endothelial cells, and interstitial fibroblasts (23, 103). All the above cell types also express components of the RAS. Pulmonary microvascular endothelial cells express high levels of ACE, contributing to the systemic regulation of BP and fluid balance (10, 105). Within the lung, AT1R and AT2R are widely expressed. AT1R is localized to VSMCs, macrophages, and stromal fibroblasts underlying the airway epithelium, whereas AT2R is expressed in the bronchial epithelium with strong expression at the brush borders (13). AT2R expression is also observed in fibroblasts, macrophages, and a subset of pulmonary endothelial cells (13). Both ACE2 and MasR are predominantly expressed by bronchial epithelial cells (75, 150), although alveolar epithelial cells, vascular endothelial cells, and VSMCs are also positive for ACE2 (59). In addition to pulmonary arterial hypertension (PAH) (30, 97), the lung RAS is involved in the pathogenesis of many other lung diseases unrelated to BP and fluid balance, which include lung infection/inflammation (74), acute lung injury/acute respiratory distress syndrome (70), COPD (60, 123), and pulmonary fibrosis (12, 137, 138).
Nicotine has been shown to increase the expression and/or activity of ACE in the lung. With the use of an ex vivo lung perfusion model, the conversion of ANG I to ANG II was found to be increased in lungs isolated from rats that were exposed to cigarette smoke on the prior day (7). This increase, however, was not sustained after repeated 10-day exposure, indicating a time- or exposure (acute vs. chronic)-dependent effect (7). In healthy human volunteers, serum ACE activity showed a significant increase immediately after smoking and returned to control level 20 min after smoking (80). Increased serum ACE levels were also found in dogs after exposure to cigarette smoke or intravenous nicotine infusion (126). In addition, nicotine and its metabolites have been shown to increase the expression and/or activity of ACE in cultured endothelial cells (94, 149).
In PAH patients, ACE expression was significantly increased in the endothelial lining of the pulmonary arteries/arterioles, areas that are normally devoid of ACE expression (in contrast to alveolar capillary endothelium, which expresses high levels of ACE) (108). The arteriole expression of ACE is consistent with the hypothesis that locally increased production of ANG II contributes to the process of pulmonary vascular remodeling in PAH. In chronic cigarette smoke-induced PAH in rats, ANG II level in the lung was increased with increased expression of ACE and decreased expression of ACE2 (60, 148). The decreased expression of ACE2 in pulmonary artery smooth muscle cells from cigarette smoke-exposed rats correlated with increased proliferation (60). In addition, losartan administration in vivo reduced ANG II level in the lung, restored ACE2 expression, and ameliorated pulmonary vascular remodeling and smoke-induced increase in right ventricular systolic pressure (60, 148). Koka et al. (81) showed that ANG II downregulates ACE2 via AT1R-mediated extracellular signal-regulated kinase (ERK)/p38 MAPK signaling pathways; thus the decreased ACE2 expression by chronic cigarette smoke exposure could be mediated by a mechanism dependent on ANG II and AT1R. Cigarette smoke exposure in mice also increases ACE expression in the lung, and this upregulation was greater in ACE2 knockout mice compared with the wild-type controls (69). In addition, ACE2 knockout mice exhibited increased inflammation with activation of MMPs and signal transducer and activator of transcription 3 following cigarette smoke exposure (69).
The association between cigarette smoking and the development of COPD is now widely accepted. Podowski et al. (111) showed that treating mice chronically exposed to cigarette smoke with losartan protected them from cigarette smoke-induced increase in lung oxidative stress levels, alveolar septal cell apoptosis, fibrosis, and emphysema development. In this study, cigarette smoke was shown to increase AT1R expression in lung parenchyma (alveolar walls), which was normalized by losartan treatment. In patients with COPD, there was a five to sixfold increase in the ratio of AT1R to AT2R in regions of marked fibrosis surrounding the bronchioles, which correlated well with reduced lung function (13). In a cigarette smoke-induced murine COPD model, administration of ANG-(1–7) (0.3 mg/kg for 2 wk) by subcutaneous infusion using osmotic pumps reduced cigarette smoke-induced lung inflammatory responses and lung fibrosis, and these effects were accompanied by increased expression of MasR and reduced NFκB signaling (150).
Maternal nicotine intake through subcutaneous osmotic minipumps has been shown to adversely affect alveolar development (with reduced radial alveolar count) and promote interstitial pulmonary fibrosis in adult male offspring rats, and these effects were accompanied by increased AT1R expression in the lung (28). The upregulation of AT1R was not observed in adult female offspring, which were protected from developmental nicotine exposure (28). In addition, AT2R was upregulated in the lung by nicotine in adult female offspring rats, which was not observed in the male offspring, indicating a gender-specific effect of nicotine (28).
LIMITATIONS OF CURRENT NICOTINE LITERATURE
Nicotine dosage and route of administration.
When comparing studies from humans, animals and culture-based assays, the major variables include nicotine dosage, route of administration, and length of exposure (Table 1). For in vivo studies, the gold standard should be serum nicotine or cotinine (metabolite of nicotine) levels (9), which were not measured in most studies (Table 1). In addition, the metabolism of nicotine differs between humans and experimental animals, which makes it difficult to extrapolate findings from animals to human smokers/e-cig users. An excellent set of guidelines on nicotine dose selection for in vivo research has been published by Matta et al. (98).
Table 1.
Nicotine dosage, route of administration, and length of exposure
| Nicotine Dosage | Route of Administration | Length of Exposure | Nicotine/Cotinine Levels | References | |
|---|---|---|---|---|---|
| Cigarette Smoke* | |||||
| Human | Acute smoking episodes | Inhalation | Over 24-h period | ND | (16) |
| 21 ± 5 Cigarettes/day | Inhalation | Nonsmoker vs. chronic smoker (23 ± 8 yr) | ND | (18) | |
| 1 Cigarette/15 min for 1 h or 2 cigarette/h for 1 day | Inhalation | 1 or 24 h | ND | (56) | |
| 5 Cigarettes for men and 3 cigarettes for women | Inhalation | 10 min | ND | (80) | |
| 5–32 Cigarettes/day | Inhalation | Nonsmoker vs. chronic smoker (12–35 yr) | 1,406 ± 333 ng/ml in smokers and 60 ± 10 ng/ml in nonsmokers (urine cotinine) | (86) | |
| 1 Cigarette during controlled administration | Inhalation | 1 Cigarette (controlled administration) + 1-h ad libitum use | 7.86–29.23 ng/ml (plasma nicotine) | (146) | |
| Rat | 5 Cigarettes/day | Inhalation chamber | 1 h/day for 1 or 10 days | ND | (7) |
| 6 Cigarettes/day | Inhalation chamber | 6 wk | ND | (11) | |
| 15 Cigarettes/30 min, twice/day | Inhalation chamber | 6 mo | ND | (60, 148) | |
| 450 mg/m3 of PM | Nose only | 2 × 1 h/day, 5 days/week for 30, 60 or 90 days | ND | (99) | |
| 8 Cigarettes/10 min, twice/day | Inhalation chamber | 8 wk | ND | (150) | |
| Mouse | Not indicated | Inhalation chamber | 4 × 30 min/day, 7 days/week for 1 to 3 wk | ND | (69) |
| 90 mg/m3 of PM | Inhalation chamber | 2 h/day, 5 days/week for 6–7 wk | ND | (111) | |
| 120 mg/m3 of PM | Inhalation chamber | 6 h/day, 4 days/week for 6 wk | ND | (133) | |
| Culture | 5 Cigarettes lit for 5 min, drawn into PBS | Culture media | 24 h | NA | (133) |
| Nicotine | |||||
| Dog | 2.5–50 μg/kg | Injection into pulmonary artery | Single injection | ND | (80) |
| 500 μg | Direct adrenal injection in vitro | 55 Injections at 100-s intervals | NA | (84) | |
| Rat | 4 μg·kg−1·min−1 | Osmotic mini-pump | 4th day of pregnancy to P10 | ND | (28, 90, 91, 96, 141–143) |
| 5 or 15 mg/kg | Subcutanous injection | Single or multiple injections | ND | (65) | |
| 15 or 48 mg·kg−1·day−1 | Osmotic minipump | 1–27 days | ND | (65) | |
| 0.1–100 pmol | Intracerebroventricular injection | Multiple injections at 3-day intervals | ND | (106) | |
| 1.5 mg·kg−1·day−1 | Subcutanous injection | Daily injection (gestational days 3 to 21) | ND | (147) | |
| Mouse | 5 mg·kg−1·day−1 | Osmotic minipump | 4 wk | ND | (22) |
| 4 mg·kg−1·day−1 | Osmotic minipump | 28 days | ND | (57) | |
| 1–5 mg·kg−1·day−1 | Osmotic minipump | 6 wk | ND | (139) | |
| Culture | 1–100 µM | Culture media | 4–48 h | NA | (43, 44) |
| 0–500 ng/ml | Culture media | 3 h | NA | (57) | |
| 0.1–10 µM | Culture media | 10 min–24 h | NA | (94, 149) | |
| 10–100 µM | Culture media | 45 min | NA | (102) | |
| 100 µM | Culture media | 10 s period | NA | (107) | |
| 50 µM | Culture media | 10 min | NA | (121) | |
| 200–600 nM | Culture media | 24 h | NA | (133) | |
| 0.01–0.1 µM | Culture media | 1 h | NA | (139) | |
| E-cigarettes | |||||
| Human | 18 mg/ml | Vaping | Once every 30 s for 10 min | 30–100 ng/ml (urine cotinine) | (24) |
| 11 mg/ml | Vaping | 7 min | ND | (41) | |
| 18 mg/ml | Vaping | Once every 30 s for 10 min | 75 ± 5 ng/ml (urine cotinine) | (48) | |
| 12 mg/ml | Vaping | 30 min/60 puffs sessions | 4.71 ± 0.91 ng/ml (plasma nicotine) | (101) | |
| 0–18 mg/ml | Vaping | Prospective 3.5-yr observational study | ND | (112) | |
| Not indicated | Vaping | Hypertensive smokers who switched to e-cig, 12-mo | ND | (113) | |
| 16–24 mg/ml | Vaping | 50 Puffs (controlled administration) + 1-h ad lib use | 1.99–22.42 ng/ml (plasma nicotine) | (146) |
NA, not applicable; ND, not determined; P, postnatal day; PM, particulate matter from cigarette smoke.
Nicotine versus cigarette smoke/e-cig.
Tobacco smoke is a toxic mixture of more than 5,000 chemicals, with potential carcinogenic, cardiovascular, and respiratory effects (129). In addition to nicotine, other components of cigarette smoke, such as carbon monoxide and polycyclic aromatic hydrocarbons, have also been implicated in the development of CVPD (1). Thus it is important to determine whether the effects of cigarette smoke on the alteration of RAS function are due to nicotine or other components of cigarette smoke.
E-cig aerosols contain fewer numbers and lower levels of most toxicants than smoke from combustible tobacco cigarettes (103a). The nicotine concentration in commercial e-liquids varies considerably, ranging from 0 to greater than 50 mg/ml (103a). Depending on device characteristics and user behavior, nicotine consumption in some e-cig users could be greater than in cigarette smokers. In addition to nicotine, e-cig contains flavorings and humectants (propylene glycol and glycerol). Although most flavorings and humectants used in e-liquids are generally recognized as safe, their potential toxicity via the inhalation route has not been carefully determined (103a). Emerging evidence suggests that oxidant chemicals, particulates and flavorings from e-cig aerosols could result in inflammation, bronchoconstriction, and airway remodeling (78, 79).
Nicotine effects and animal behavior.
To date, virtually all data available on the effects of nicotine on cardiopulmonary function in experimental animals are based on experimenter-administered nicotine. The potential differences in experimenter-administered nicotine versus self-administered nicotine is worthy of consideration in light of the report by Donny et al. (33), which showed increased plasma catecholamine levels in rats receiving response-independent nicotine infusion (experimenter-administered nicotine) but not in rats with nicotine self-administration. This study indicates that nicotine’s acute effects can be powerfully modified by the contingency relationship between drug administration and animal behavior (33). Future studies using self-administration models in laboratory animals may yield findings with greater relevance to human smokers.
CONCLUSIONS
The literature discussed in this review strongly supports a link between nicotine and the RAS, i.e., nicotine impairs RAS homeostasis in multiple organ systems, contributing to the development of CVPD. As shown in Fig. 1, nicotine interacts with many components of the RAS. In the ACE/ANG II/AT1R arm, nicotine has been shown to increase the expression and/or activity of renin, ACE and AT1R, whereas in the compensatory ACE2/ANG-(1–7)/MasR arm, nicotine downregulates the expression and/or activity of ACE2 and AT2R. Nicotine-induced imbalance of the two arms of the RAS is likely responsible for many of the adverse actions of nicotine in the development and exacerbation of CVPD, including hypertension, dysregulated cardiac remodeling, vascular dysfunction, and chronic lung diseases. Tissue-specific RAS and the effects of cigarette smoke and nicotine on RAS components are summarized in Table 2. Because of the complexity of the RAS (Fig. 1), the effects of nicotine could be beyond what we have discussed in this review. For example, the effect of nicotine on the ANG A/alamandine/Mas-related G protein-coupled receptor D axis, in which ACE2 also plays a critical role, is completely unknown.
Table 2.
Tissue-specific RAS components and their major roles
| RAS | Heart and Vasculature | Brain and ANS | Lung |
|---|---|---|---|
| ACE | Vasoconstrictive (18), CS↑ (18) | Prohypertensive (4) | Increased expression and/or activity in PAH (30, 108), CS↑ (7, 60, 69, 80, 126), N↑ (126) |
| N↑ expression in glial cells (43) | |||
| ACE2 | Vasodilatory (128), CS↓ (60) | Antihypertensive (125) | Anti-inflammatory (69, 138) |
| Anti-inflammatory (125) | Antifibrotic (138) | ||
| N↓ expression in both neurons and glial cells (43, 44) | Decreased expression in PAH, CS↓ (60, 148) | ||
| ANG II | Prohypertrophic (22), N↑ (22) | Prohypertensive (4) | Proproliferative for PA-SMC (30) |
| Vasoconstrictive (8, 141, 143), N↑ (141, 143) | Increased level in PAH (30), CS↑ (60, 148) | ||
| Prohypertensive (125) | |||
| AT1R | Prohypertrophic (72) | Prohypertensive (4) | Proinflammatory (111), CS↑ (111) |
| Profibrotic (72) | Sympathostimulatory (102, 106, 107) | Profibrotic (111, 137), N↑ (28) | |
| Vasoconstrictive (8, 134), N↑ (141, 143) | N↑ expression in both neurons and glial cells (43, 44) | ||
| N↑ expression in the brain (96) | |||
| AT2R | Antihypertrophic (114) | Sympathoinhibitory (51, 52, 102) | Anti-inflammatory (12) |
| Antifibrotic (114) | N↓ expression in the brain (91, 96) | Antifibrotic (12) | |
| Vasodilatory (8, 72, 128, 136), N↓ (141, 143) | Decreased expression in COPD (13) | ||
| ANG-(1–7) | Antihypertrophic (72) | Antihypertensive (145) | Anti-inflammatory (150) |
| Antifibrotic (72) | Reduces norepinephrine release and TH expression (145) | Antifibrotic (150) | |
| Vasodilatory (72, 128, 136) | |||
| MasR | Vasodilatory (72) | Increases baroreflex activation (145) | Anti-inflammatory (150) |
| Antifibrotic (150) |
ACE, angiotensin-converting enzyme; AT1R and AT2R: ANG II type 1 and type 2 receptor; MasR, Mas receptor; ANS, autonomic nervous system; COPD, chronic obstructive pulmonary disease; CS↑, upregulated by cigarette smoke; CS↓, downregulated by cigarette smoke; N↑, upregulated by nicotine; N↓, downregulated by nicotine; PA, pulmonary artery; PAH, pulmonary arterial hypertension; RAS, renin-angiotensin system; SMA, smooth muscle cells; TH, tyrosine hydroxylase.
Perspectives and Significance
This review summarizes our current knowledge of the interaction between nicotine and the RAS, one of the most important regulatory systems of cardiopulmonary function in health and disease. The current review aims to raise the awareness of the harmful effects of nicotine and to stimulate continued nicotine research. Many questions remain to be answered. What are the mechanisms by which nicotine alters the RAS in different organ systems? Which nicotinic receptors and/or intracellular signaling pathways are involved in the modification of RAS by nicotine? What are the effects of continued nicotine intake on the therapeutic actions of antihypertensive drugs (ACE inhibitors and/or angiotensin receptor blockers) in hypertensive patients? With the emerging popularity of e-cig among youth and young adults (82), age-, developmental- and gender-specific effects of nicotine are also important areas of nicotine investigation.
GRANTS
This work was supported by the National Heart, Lung, and Blood Institute Grant R01-HL-135635 and the National Institute of General Medical Sciences Grant P30-GM-106392.
DISCLOSURES
No conflicts of interest, financial or otherwise, are declared by the authors.
AUTHOR CONTRIBUTIONS
J.M.O., R.M.F., E.L., and X.Y. prepared figures; J.M.O., R.M.F., E.L., and X.Y. drafted manuscript; J.M.O., R.M.F., J.D.G., E.L., and X.Y. edited and revised manuscript; J.M.O., R.M.F., J.D.G., E.L., and X.Y. approved final version of manuscript.
ACKNOWLEDGMENTS
Present address of E. Lazartigues: Dept.of Pharmacology and Experimental Therapeutics, Louisiana State Univ. Health Sciences Center, New Orleans, LA 70112 (e-mail: ude.cshusl@razale, ude.cshusl@razale).
REFERENCES
Articles from American Journal of Physiology - Regulatory, Integrative and Comparative Physiology are provided here courtesy of American Physiological Society
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