Effect of high‐altitude trekking on blood pressure and on asymmetric dimethylarginine and isoprostane production: Results from a Mount Ararat expedition

Abstract

The study aimed at exploring the mechanisms behind blood pressure and heart rate changes upon acute altitude exposure utilizing urinary excretion of biochemical factors involved in cardiovascular regulation. The study was conducted on 12 lowlander native male mountain climbers, living at sea level, exposed to altitudes ranging from 1800 to 5147 m above sea level over 4 days, during their ascent to Mount Ararat (Turkey). Blood pressure (measured by oscillometric method), heart rate, and blood oxygen saturation (SpO₂) were recorded at rest (on awakening before food intake), in hypoxic conditions at 4200 m and at sea level before and after the altitude expedition. In the same study conditions (ie before‐during‐after the expedition), first‐voided urinary samples were collected and assayed for 8‐iso‐prostaglandin F_2α (8‐iso‐PGF_2α) and asymmetric dimethylarginine (ADMA) determination. Heart rate, and systolic and diastolic blood pressures were higher (P < .05) at high altitude than at the sea level. Furthermore, both urinary 8‐iso‐PGF_2α and ADMA were significantly elevated (P < .01) at high altitude and returned to normal levels soon after returning to sea level. A 4‐day exposure to high‐altitude hypoxia induced a temporary increase in blood pressure and heart rate, confirming previous findings. Blood pressure increase at high altitude was associated with significantly enhanced production of biochemical mediators such as 8‐iso‐PGF2α, catecholamines, and ADMA, although we could not demonstrate a direct link between these parallel significant changes probably due to the forcefully limited sample size of our study, carried out in challenging environmental conditions at very high altitude.

1. INTRODUCTION

Acute high‐altitude exposure triggers several adaptive changes through various physiological mechanisms, represented by modifications in blood composition and in neural, cardiovascular, renal, metabolic, respiratory, and reproductive functions. ¹ , ² , ³ , ⁴ , ⁵ , ⁶ , ⁷ , ⁸

Recent data indicate that acute exposure to high altitude may influence various mechanisms of blood pressure (BP) control (ie, neural central and reflex control of sympathetic activity, arterial elasticity, endothelial function, blood viscosity), resulting in BP and heart rate (HR) increases in healthy and in hypertensive patients, which persist after acclimatization. ² , ⁴ , ⁹ , ¹⁰ , ¹¹ , ¹²

By exploring changes in conventional BP and in 24‐hours ambulatory BP when moving from sea level to high altitude (HIGHCARE‐Himalaya study), Parati et al showed that the exposure to progressively higher altitudes was associated with a progressive and marked increase in systolic and diastolic ambulatory BP levels (SBP/DBP), more evident than the increase in conventional BP, paralleled by an increase in plasma catecholamine concentration. This increase was accompanied by a reduction in nocturnal BP dipping, possibly due to a further nocturnal reduction of SpO₂. This phenomenon is accompanied by an additional increase in sympathetic activity, which might be related to the frequent occurrence of central apneas during sleep at high altitude. ³ These findings are in agreement with those by Reeves et al, ¹³ who also showed that exposure to high‐altitude hypoxia increases both systemic arterial pressure and sympathetic activity, concomitantly with the increase in norepinephrine (NE) levels, related to an increased alpha adrenergic‐mediated vascular tone.

Recent literature highlights the role of nitric oxide (NO) in the mechanisms that regulate both pulmonary ¹⁵ and systemic blood pressure. ¹⁶ , ¹⁷ In this regard, a role for asymmetric dimethylarginine (ADMA), an inhibitor of NO synthesis favoring vascular vasoconstriction ¹⁸ , ¹⁹ and renal sodium excretion or inhibition, ²⁰ , ²² has been suggested, supporting the observed blood pressure increase.

The BP adaptation to high altitude is characterized by changes related to the duration of exposure: Over the first minutes or hours of hypoxia exposure, BP remains fundamentally unchanged due to the combination of a direct vasodilatory effect by hypoxia with a chemoreflex‐induced sympathetic activation responsible for reflex vasoconstriction. ⁹ After this first phase, activation of pressor mechanisms prevails, leading to increased BP. ¹⁴ Only after a longer period of high‐altitude sojourn, these mechanisms may be partially inhibited, due to blood oxygen concentration rising again toward baseline levels, through the acclimatization process and hematocrit increase. ²³

1.1. Asymmetric dimethylarginine (ADMA)

Asymmetric dimethylarginine is metabolized by dimethylarginine dimethylaminohydrolases (DDAHs) to citrulline and dimethylamine (DMA) and is partly excreted unchanged via the kidney. ²⁴ A small part of ADMA is cleared by urinary excretion, but it has been estimated that more than 80% is metabolized/hydrolyzed by intracellular dimethylarginine dimethylaminohydrolase (DDAH). ²⁵ Elevated ADMA levels were reported in human hypertension and in diverse animal models of hypertension. ²⁶ , ²⁷

In recent years, it has been speculated that the development and progression of arterial hypertension are, at least partially, related to ADMA‐induced NO‐ROS imbalance, ²⁶ , ²⁸ and small animal models have shown that restoration of that balance could prevent the development of hypertension. ²⁹

By inhibiting NO formation, ³⁰ ADMA causes endothelial dysfunction, vasoconstriction, elevation of BP, and aggravation of experimental atherosclerosis. ²⁶ Furthermore, there is evidence that both hypoxia and exaggerated BP response to exercise can be related to increased ADMA production. ³¹

1.2. 8‐iso‐PGF_2α

The 8‐iso‐PGF2α and 8‐iso‐prostaglandin E2 (8‐iso‐PGE2) are powerful vasoconstrictors, ³² , ³³ and it has been suggested that their direct vasoconstrictive effect, combined with their potentiating effect, at subthreshold concentrations, on the vasoconstriction induced by NE or angiotensin II (Ag II), might be of pathophysiological relevance in cardiovascular diseases. ³⁴ This has been suggested by Sametz and colleagues who have shown that infusion, in the isolated rabbit ear, of 8‐iso‐prostaglandin E2 and 8‐iso‐prostaglandin F2alpha, amplified significantly the vasoconstriction induced by norepinephrine or angiotensin II.

It has also been suggested that high‐altitude exposure could lead to increased plasma and urinary levels of 8‐iso‐prostaglandin F2α (8‐iso‐PGF2α). ³⁵ , ³⁶ An increased serum 8‐iso‐PGF2α in hypertensive double‐transgenic rats (dTGR) exposed to acute hypoxia has indeed been described. ³⁷ 8‐Iso‐PGF2α is an isomer of classical prostaglandins that is principally produced by oxidative stress–mediated pathways and is used as a diagnostic biomarker for evaluating lipid peroxidation, in vivo. ³⁸

1.3. Aim of our study

The aim of this study was to evaluate the changes in BP and those in urinary excretion of 8‐iso‐PGF_2α, ADMA, epinephrine (E), and NE in non‐acclimatized patients acutely exposed to hypobaric hypoxia at altitude. Specifically, the focus of the study was to investigate the role of ADMA production in the regulation of vascular tone and BP.

2. MATERIALS AND METHODS

2.1. Participants, ascent/descent altimetric plan

The Declaration of Helsinki ethical principles for human medical research have been adhered to, and the study was approved by the Ethical Committee of “G. D’Annunzio” University, Chieti, Italy.

The patients included in this study were 12 healthy male lowlanders (age: 42 ± 11.0 years; BMI: 26 ± 2.5 Kg/m²) non‐acclimatized to altitude, non‐smokers, patients with no history of chronic diseases (such as hypoxic pathologies and chronic hypertension), and patients with experience of mountain climbing but no recent exposure to high altitude over the 6 months prior to this study (Table 1). The study protocol included data collection during ascent to Mount Ararat (altitude of 5147 m, Turkey), and after return to sea level, which occurred over 4 days, during which participants remained at an altitude ranging from 1800 to 5147 m. On the first day, participants ascended by car from Dogubayazit (1800 m) to Cevirme village located at the foot of Mount Ararat at 2200 m, from where they started trekking to reach camp 1 at an altitude of 3200 m where the group spent the night (day 1). On the second day of climb, participants reached camp 2, at an altitude of 4200 m, where they spent the night (day 2). On the third day, the group stayed at an altitude of 4200 for acclimatization (day 3), and on the fourth day, they reached the summit of Mount Ararat at an altitude of 5147 m and then descended toward the altitude of 2200 m, near Cevirme village, from where they were brought by car down to Dogubayazit at 1800 m (day 4). The phases of the ascent are described in Figure 1.

TABLE 1.

Description of the patients participating to the study

Age	Weight (Kg)	Height (cm)	BMI
50	73	168	26
34	71	178	22
48	68	169	24
51	67	165	25
34	87	168	31
22	67	164	25
55	91	176	29
42	90	183	27
57	94	180	29
27	77	174	25
45	82	178	26
40	82	182	25

FIGURE 1.

Ararat expedition altimetric profile, experimental evaluation phases in the expedition timeline, and SpO₂ response to high altitude

2.2. Study protocol and measurements

Cardiovascular measurements (BP, HR, and SpO₂) and urine collection for ADMA, 8‐iso‐PGF2α, and catecholamines (NE and E) determination were performed at rest, on awakening, around 9 AM, and fasting, in three different moments: at sea level 1 day before the expedition (sl^B); at Camp 2 at 4200 m on the third trekking day (Camp2⁴²⁰⁰); and at sea level the day after the expedition (sl^A).

2.3. BP, HR, and SpO₂ determination

Blood pressure and HR were recorded by an oscillometric electronic devices (Omron M6 [HEM‐7001‐E], Omron Healthcare), validated according to the International Protocol of the European Society of Hypertension, ³³ while SpO₂ was evaluated by a finger pulse oximeter (503 OXY‐5 GIMA, Gima S.p.A.). The participants were requested to rest in the sitting position for 5 minutes before measurements, without crossing their legs, and to remain silent before and during the measurements that were made using the same arm held in a horizontal position at heart level with a support bracket. The average of 2 BP measurements taken at 1‐min interval was stored in the study Case Report Form (CRF).

2.4. Urine collection, preparation, and 8‐iso‐PGF2α, NE, E, and ADMA assessment

The obtained first‐voided urinary samples were immediately stored and transferred in ice (−20°C) and subsequently analyzed. Specifically, as described in study protocol and measurements, the urinary samples were collected at sl^B, at Camp2⁴²⁰⁰, and at sl^A. Urine analysis was carried out in Italy in the Department of Pharmacy Laboratory, “G. d’Annunzio” University of Chieti, where 1 mL urine samples were stirred for 1 minutes using a vortex mixer, centrifuged at 13 000 g for 10 minutes to remove sediments, and finally filtered through Millipore 0.25‐µm nylon filters. This was followed by radioimmunoassay for 8‐iso‐PGF2α assessment; HPLC‐EC essay for NE and E assessment; and HPLC‐UV for ADMA and creatinine assessment. ³⁰ , ³¹ , ³² , ³³ , ³⁴ The detailed protocols are described below.

2.5. Radioimmunoassay 8‐iso‐PGF_2α determination

25 µL urine aliquots were 10‐fold diluted in 25 µmol/L phosphate buffer, and 8‐iso‐PGF_2α levels were measured by radioimmunoassay (RIA), as previously described. ³⁹ Briefly, specific anti‐8‐iso‐PGF_2α serum was developed in the rabbit; its cross‐reactivity with 8‐iso‐PGE₂ was 7.7%, whereas with other prostanoids, it was <0.3%. ⁴⁰ The detection limit of the assay method is 6 pg/mL, and the IC₅₀ is 39.8 pg/mL. The intraassay and interassay coefficients of variation are ± 2.0% and ± 2.9%, at the lowest concentration of standard (2 pg/mL), and ± 3.7% and ± 9.8%, at the highest concentration of standard (250 pg/mL).

2.6. High‐performance liquid chromatography (HPLC)—UV ADMA and creatinine determination

Urinary ADMA and creatinine levels were determined as previously reported. ⁴¹ , ⁴² , ⁴³ The HPLC apparatus consisted of a Jasco PU‐2080 chromatographic pump and a Jasco MD‐2010 Plus absorbance detector. Integration was performed by Jasco Borwin Chromatography software, version 1.5. The chromatographic separation was performed by isocratic elution on GraceSmart reverse‐phase column (C18, 150 mm × 4.6 mm i.d., 5 µm). The mobile phase was (1:99, v/v) acetonitrile and 25 mmol/L pH 5.00 phosphate buffer containing octane sulfonic acid 10mM and triethylamine 0.03% v/v. The flow rate was 1.3 mL/min, and the samples were manually injected through a 20‐µL loop. ADMA analyses were performed by injecting 20 µL urinary aliquots in the HPLC and monitoring absorbance at 200 nm. On the other hand, creatinine analyses were performed by 200‐fold diluting urinary samples before injection and monitoring absorbance at 235 nm. ADMA and creatinine peaks were identified by comparison with pure standard retention time, while their concentrations in the urinary samples were calculated by linear regression curve (y = bx + m) obtained with a standard. The urinary recovery was satisfactory with RSD %< 10. The standard stock solution of ADMA at 2 mg/mL was prepared in bidistilled water containing 0.004% EDTA and 0.010% sodium bisulfate, while standard stock solution of creatinine (2 mg/mL) was purchased from Alexis Biochemicals. The stock solutions were stored at 4°C. Work solutions (20‐200 µg/mL) were daily obtained, progressively diluting stock solutions in the mobile phase.

2.7. HPLC norepinephrine (NE) and epinephrine (E)

Norepinephrine and E levels were analyzed through an HPLC apparatus consisting of a Jasco PU‐2080 chromatographic pump and an ESA Coulochem III coulometric detector, equipped with a microdialysis cell (ESA‐5014b) porous graphite working electrode and a solid‐state palladium reference electrode. The experimental conditions for biogenic amine identification and quantification were selected as follows. The analytical cell was set at − 0.150 V for detector 1 and + 0.300 V for detector 2, with a range of 100 nA. The chromatograms were monitored at the analytical detector 2. Integration was performed by Jasco Borwin Chromatography software version 1.5. The chromatographic separation was performed by isocratic elution on a Phenomenex Kinetex reverse‐phase column (C18, 150 × 4.6 mm i.d., 2.6 µm). As regards the separation of NE and E, the mobile phase was (10:90, v/v) acetonitrile and 75 mmol/L pH 3.00 phosphate buffer containing octane sulfonic acid 1.8 mmol/L, EDTA 30 µmol/L, and triethylamine 0.015% v/v. The flow rate was 0.6 mL/min, and the samples were manually injected through a 20‐µL loop. Neurotransmitter peaks were identified by comparison with the retention time of pure standard. Neurotransmitter concentrations in the samples were calculated by linear regression curve (y = bx + m) obtained with a standard. Neither internal nor external standard was necessary for neurotransmitter quantification in the hypothalamus homogenate, and all tests performed for method validation yielded results in accordance with limits indicated in official guidelines for applicability in laboratory trials. The standard stock solutions of NE and E at 2 mg/mL were prepared in bidistilled water containing 0.004% EDTA and 0.010% sodium bisulfite. The stock solutions were stored at 4°C. Work solutions (1.25‐20.00 ng/mL) were obtained daily by progressively diluting the stock solutions in the mobile phase.

2.8. Statistical analysis

Data are presented as means ± standard deviation (SD) and analyzed by repeated‐measures ANOVA using unstructured variance‐covariance matrix between observations of the same patient and adjusting for age and BMI. ANOVA models were fitted to evaluate the relationship between the altitude levels and (a) each BP or HR indicator (SBP, DBP, HR) and (b) each urinary biochemical parameter considered (SpO₂, urinary 8‐iso‐PGF2α, ADMA, NE, E). In any model, for each altitude level, the least square mean (LS mean), with its standard error (SE), was calculated. LS means comparisons (Camp2⁴²⁰⁰ vs sl^B and Camp2⁴²⁰⁰ vs sl^A) were performed taking into account the multiple‐testing Dunnett adjustment. The relations among changes in SpO2, urinary 8‐iso‐PGF2α, ADMA, NE, E with BP were explored by multivariate mixed regression analysis (separately for systolic and diastolic BP) using compound symmetry as correlation matrix between observations of the same patient and adjusting for age and BMI. All analyses were performed using the Statistical Analysis System software (version 9.4; SAS Institute). Statistical significance was set at the 0.05 level.

3. RESULTS

During the expedition, there were no medical problems related to altitude. Furthermore, the experimental patients, given the aim of our study for the study, did not make use of acetazolamide (Diamox). Although the STAR Data Reporting Guidelines for Clinical High‐Altitude Research were published after our expedition, we can say “a posteriori” that the study was conducted in accordance with them. ⁴⁴

While SpO₂ measured at Camp2 was significantly lower than at sl^B and at sl^A (P < .0001 and P < 0. 0001 for both individual comparisons) (Figure 2A), HR (Figure 2B), all BP values (Figure 3) and urinary concentrations of biochemical parameters were significantly higher at Camp2⁴²⁰⁰ than at sl^B and sl^A (Figure 4A‐D).

FIGURE 2.

A, SpO₂ measurements in the three different study conditions: at sea level before the expedition (sl^B); to Camp 2 at 4200 m on the third trekking day (Camp2⁴²⁰⁰), and at sea level after the expedition (sl^A). B, HR (beats/min) measured at sl^B, at sl^A, and at the Camp2⁴²⁰⁰. Least squared means of HR (betas/min) from an ANOVA model including age and BMI. Data are presented as means ± standard deviation (SD)

FIGURE 3.

Systolic and diastolic BP (mm Hg) measured at sl^B, at sl^A, and at the Camp2⁴²⁰⁰. Least squared means of systolic and diastolic BP (mm Hg) from an ANOVA model including age and BMI. Data are presented as means ± standard deviation (SD)

FIGURE 4.

A, Urinary 8‐iso‐prostaglandin F_2α (8‐iso‐PGF_2α) excretion expressed as pg/mg creatinine measured at sl^B, at sl^A, and at the Camp2⁴²⁰⁰. Least squared means of Urinary 8‐iso‐prostaglandin F2α (8‐iso‐PGF2α) excretion, expressed as pg/mg creatinine, from an ANOVA model including age and BMI. B, Urinary asymmetric dimethylarginine (ADMA) excretion expressed as µg/mg creatinine measured at sl^B, at sl^A, and at the Camp2⁴²⁰⁰. Least squared means of urinary asymmetric dimethylarginine (ADMA) excretion, expressed as µg/mg creatinine, from an ANOVA model including age and BMI. C, Urinary NE excretion measured at sl^B, at sl^A, and at the Camp2⁴²⁰⁰. Least squared means of urinary NE excretion, expressed as ng/mg creatinine, from an ANOVA model including age and BMI. D, Urinary E excretion measured at sl^B, at sl^A, and at the Camp2⁴²⁰⁰. Least squared means of urinary E excretion, expressed as ng/mg creatinine, from an ANOVA model including age and BMI. Data are presented as means ± standard deviation (SD)

In particular, urinary 8‐iso‐PGF_2α (Figure 4A) and ADMA (Figure 4B) (indexed to creatinine as picograms or nanograms per milligram creatinine, respectively) were significantly higher at the Camp2⁴²⁰⁰, than in normoxic conditions at sl^B or at sl^A, and a similar behavior characterized also urinary NE and E (Figure 4C and Figure 4D). Since the creatinine excretion was unchanged between sea level and altitude, we are confident that the excretion of metabolites was real and not altered by changes in renal function.

Although all changes from sea level to high altitude were highly significant in univariate analysis, when we explored the possible relation between BP changes and changes in the biochemical parameters assessed in our study through different multivariate analysis models, neither biochemical variable nor their changes were significantly related to systolic and diastolic BP values nor to their changes, respectively (Table 2), although we cannot exclude the occurrence of a nonlinear relationship between these variables.

TABLE 2.

P‐values from mixed regression models

Variable	Systolic BP a	Diastolic BP a
SpO2	0.5712	0.7001
8‐iso‐PGF2α	0.3665	0.3667
ADMA	0.9035	0.2861
NE	0.1583	0.1327
E	0.7224	0.7762

^aModel adjusted for age, BMI, and altitude.

4. DISCUSSION

Previous studies on the BP changes occurring under acute exposure to hypobaric hypoxia at altitude carried out up to the end of the 20th century provided discordant results. While some investigators found a small increase in BP with hypoxia exposure, others reported a small reduction or even no changes. ⁴⁵ Vogel et al concluded that acute short‐lasting 1‐4 days hypoxia exposure has little influence on systemic BP in humans. ⁴⁶ Similarly, Rostrup concluded on the absence of BP changes in response to altitude‐related hypoxia. ⁴⁷ On the other hand, Bestle et al reported that several days of exposure to altitude‐related hypoxia induced an elevation of BP compared with the sea level. ⁴⁸ Somehow paradoxically, a group of researchers, mostly from the former Soviet Union, have been suggesting since decades ago the possibility of an antihypertensive effect by exposure to intermittent hypoxia, with intermittent hypoxic breathing programs being proposed as a type of non‐pharmacologic therapy for patients with arterial hypertension. ⁴⁹ , ⁵⁰ Recently, a more systematic approach to the investigation of the effects of acute exposure to high altitude on BP, based not only on conventional BP measurements, but also on ambulatory BP monitoring over 24 hours, has been implemented in the framework of the HIGHCARE (HIGH altitude CArdiovascular REsearch) studies by Parati et al, ² , ³ , ⁴ showing that acute exposure to high‐altitude hypoxia, after at least 7 hours, is indeed characterized by a systematic and significant increase in BP over 24 hours, in particular at night, an increase which is more evident than the increase in conventional BP measured at rest.

In our study, characterized by carefully standardized conventional BP measurements at rest, we confirm that acute exposure to high‐altitude hypoxia is associated with an acute increase in systolic and diastolic BP (Figure 3) and in HR (Figure 2B), accompanied by an increase in urinary catecholamines further supporting the role in this regard of an increased sympathetic activity. ³ , ¹³

Another interesting result of our study is that the increase in average BP and HR found in our group at altitude was accompanied also by a significant increase in average urinary levels of 8‐iso‐PGF_2α (Figure 4A). The results of our study thus provide further support to an involvement of 8‐iso‐PGF_2α in the complex physiological mechanisms that relate altitude, oxidative stress, hypertension, and vasal activity, in pathophysiological conditions with a possible direct effect on the regulation of vascular tone through thromboxane receptor activation. ⁵¹

In our study, we also provide the demonstration of significantly increased levels of urinary ADMA, which indicate its possible involvement in the cardiovascular changes induced by acute high‐altitude exposure. Indeed, changes in sympathetic activity after renal denervation associated with simultaneous changes in plasma levels of ADMA might suggest that the sympathetic nervous system exerts a vital role in modulating circulating levels of ADMA. ⁵² However, renal denervation may simultaneously affect both ADMA and sympathetic nerve traffic, without any causal relationship between them. On the other hand, studies on patients without renal denervation suggest that sympathetic activity may accompany elevation of plasma ADMA concentrations, although these observations cannot clarify how sympathetic activity can modulate ADMA levels. ⁵³

In this regard, it is interesting to note that exposure to high altitude is associated with an increase in sympathetic activity. ³ It has also been shown that ADMA plasma levels are elevated in essential hypertension, a condition characterized both by an increase in sympathetic activity and by an increased oxidative stress. ⁵⁴ Elevated BP, and increased plasma and urinary ADMA levels have been observed in Sprague Dawley rats treated with angiotensin II and in Dahl salt–sensitive rats fed with a high‐salt diet. ³⁰ , ⁵⁵ Furthermore, angiotensin II–treated rats show both increased plasma ADMA and increased urinary isoprostane, while acute angiotensin II infusion increases oxidative stress in hypertensive humans enhancing plasma isoprostane levels. ⁵⁶ It has also been found that hypoxia‐induced vascular ET‐1 release ⁵⁷ could increase vascular resistance, by stimulating ADMA production. ⁵⁸ The human kidney is responsible for ADMA and symmetrical dimethylarginine (SDMA) urinary excretion, clearing a substantial amount of ADMA by metabolic degradation, probably by DDAH. ⁵⁹ A rat chronic hypoxia–induced pulmonary hypertension model is associated with increased pulmonary concentrations of ADMA. Moreover, pulmonary hypertensive rats exhibit reduced pulmonary expression and activity of the ADMA‐metabolizing enzyme DDAH I. The decreased DDAH I and increased ADMA concentrations may therefore contribute to pulmonary hypertension via the competitive inhibition of pulmonary NOS enzymes. ¹⁵ Furthermore, it has been reported that under chronic hypoxia, ADMA increases, whereas DDAH activity decreases. ⁶⁰ Therefore, it is reasonable to think that the elevated urinary values of ADMA obtained are likely due to an increased urinary ADMA excretion as a result of systemic accumulation by its overproduction and reduced clearance.

Finally, other researchers have hypothesized that both high altitude–induced BP dysregulation and isoprostane and ADMA overproduction could be related to the activation of several biochemical pathways, such as the renin‐angiotensin and ET‐1 systems, which have been causally related to hypoxia, hypertension, and oxidative stress both in animals and in humans. ⁵⁶ , ⁶¹

Asymmetric dimethylarginine is not only an inhibitor of NO synthesis modulating pulmonary arterial pressure, ¹⁵ but also a robust marker of endothelial dysfunction, ¹⁶ whose levels are higher in patients with essential hypertension ⁵⁴ compared with normotensive individuals (P = .031). Along the same line, in 2004, Kielstein et al ¹⁷ showed that a systemic infusion of ADMA led to an increase in systemic vascular resistance and BP in a dose‐dependent manner.

The pathophysiological mechanisms of ADMA‐related hypertension therefore include its role in the regulation of vascular tone and BP, ⁶² , ⁶³ with inhibition of endothelial NOS activity responsible for vasoconstriction ¹⁸ , ¹⁹ and reduction in renal NO synthesis, and inhibition of the renal excretion of sodium by. ²⁰ , ²¹ , ²² All these mechanisms are likely to be involved in the parallel increase in ADMA and BP that we have observed under acute exposure to very high altitude.

4.1. Study merits and limitations

Our study has the merit of having jointly and systematically explored not only the cardiovascular changes induced by acute exposure to hypobaric hypoxia at altitude but also the changes in a number of biochemical variables. These variables are known to play an important role in cardiovascular regulation, thus providing additional information on possible mechanisms responsible for the cardiovascular changes observed with acute altitude exposure. It also has a limitation, however, that is the relatively small number of patients recruited and available to participate to data collection in the challenging environmental conditions of our study, a feature common to a number of expeditions organized in the past to explore the biological effects of acute exposure to very high‐altitude settings. This limited sample size is the most likely explanation of our inability to provide significant evidence of a linear relationship between BP and humoral changes at altitude. Nevertheless, our study provides novel and solid information on the occurrence of significant changes both in BP and in a number of biochemical parameters in our patients when acutely exposed from sea level to an altitude of 4200 m, suggesting the possibility of a nonlinear association between them.

Although our study cannot clarify the type of relation occurring between these hemodynamic and biochemical changes, it paves the way for further investigations, which might address the issue explored in our study more in depth, by collecting additional information in a larger number of individuals.

5. CONCLUSIONS AND PERSPECTIVES

We provide additional evidence that an acute and short exposure to high‐altitude hypoxia is associated with a HR and BP rise. We also demonstrate that these increases are paralleled by increased 8‐iso‐PGF_2α, catecholamines, and ADMA urinary excretion, although the type of relation characterizing hemodynamic and biochemical changes still needs further evaluation. Our present findings highlight the concomitant occurrence of BP rise and of significant changes in oxidative stress markers, in markers of endothelial dysfunction and in the release of vasoconstrictive prostaglandins, all accompanying sympathetic activation, in response to hypobaric hypoxia exposure at high altitude. These results emphasize the need of further studies aimed at defining the possible role of ADMA and 8‐iso‐PGF_2α as new predictive biomarkers of cardiovascular changes induced by exposure to hypoxic conditions.

Although humans have systems to counter hypoxia stress, ⁶⁴ , ⁶⁵ when considering the need of adequately protecting patients with preexisting cardiovascular conditions who decide to expose themselves to hypobaric hypoxia at altitude, as very recently highlighted in an international position paper on this issue driven by our group, ⁶⁶ our results may help identifying novel pharmacologic targets to antagonize, the oxidative stress, and BP increase induced by high‐altitude exposure.

CONFLICTS OF INTEREST

The authors declare that they have no conflicts of interest in relation to this study data.

AUTHORS' CONTRIBUTION

VV, C.F, and G.P conceived and designed the study; VV, C.F, and D.S performed the experiments; VV, CF, DS, A.Z, and G.P analyzed the data; VV, CF, DS, AZ, SB, GO, L.B, and G.P interpreted results of experiments; D.S and A.Z prepared the figures; VV, CF, D.S, and G.P drafted the manuscript; VV, CF, DS, AZ, SB, GO, L.B, and G.P edited and revised the manuscript; VV, CF, DS, AZ, SB, GO, L.B, and G.P approved the final version of the manuscript.

Verratti V, Ferrante C, Soranna D, et al. Effect of high‐altitude trekking on blood pressure and on asymmetric dimethylarginine and isoprostane production: Results from a Mount Ararat expedition. J Clin Hypertens. 2020;22:1494–1503. 10.1111/jch.13961