Blood pressure decrease in elderly after isometric training : does lactate play a role ?

This study aimed to investigate the effect of isometric handgrip (IHG) training on the blood pressure (BP) reduction in prehypertensive and hypertensive elderly people, and the possible role of lactate and redox balance. Thirty-three older (75.3±1.3 years old) were allocated to a non-exercise control (CG, n=11), prehypertensive (PHG, n=10), and hypertensive (HG, n=12) groups. PHG and HG performed a total of 8 sets of 1-min bilateral contractions at 30% maximal voluntary isometric contraction, each separated by 1-minute rest-pause. IHG training was performed for 8-week, 3 times a week on non-consecutive days. Systolic BP (SBP) and Research, Society and Development, v. 9, n. 9, e655997433, 2020 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v9i9.7433 4 heart rate (HR) decreased post-training on PHG (-10 mm Hg; -5 bpm) and HG (-16 mm Hg; 9 bpm), respectively. Diastolic BP (DBP) decreased for HG only (-9 mm Hg) (P < 0.05). In addition, the decrease in BP occurred in parallel to a better redox balance and increased bioavailability of nitric oxide in PHG and HG (P < 0.05). Also, the variables that most present association to SBP decrease were capillary blood lactate concentration and muscle strength (P < 0.05). In summary, IHG training may be practical in improving clinical status of prehypertensive and hypertensive patients, by improving BP control, NObioavailability and redox balance. Further studies are required to elucidate the pathways of lactate concentration in blood-flow during exercise.

4 heart rate (HR) decreased post-training on PHG (-10 mm Hg; -5 bpm) and HG (-16 mm Hg; -9 bpm), respectively. Diastolic BP (DBP) decreased for HG only (-9 mm Hg) (P < 0.05). In addition, the decrease in BP occurred in parallel to a better redox balance and increased bioavailability of nitric oxide in PHG and HG (P < 0.05). Also, the variables that most present association to SBP decrease were capillary blood lactate concentration and muscle strength (P < 0.05). In summary, IHG training may be practical in improving clinical status of prehypertensive and hypertensive patients, by improving BP control, NO-bioavailability and redox balance. Further studies are required to elucidate the pathways of lactate concentration in blood-flow during exercise.
Until now, it has been suggested that changes in the autonomic function (Taylor et al., 2003) and oxidative stress (Peters et al., 2006) may be mechanisms involved in BP control in hypertensive adults and older after IHG training. Currently it was demonstrated that oxidative stress has a crucial role in the pathogenesis of hypertension (Baradaran, Nasri, & Rafieian-Kopaei, 2014). It is well known that exercise positively alters the redox status of cells and tissues, possibly due to increased antioxidant defenses and nitric oxide (NO-) (de Sousa et al., 2017). However, specifically for isometric exercise, it was demonstrated that after an acute IHG session leading to BP reduction in hypertensive elderly people, no association was observed with NO- (Souza et al., 2018).
On the other hand, muscle contractions can elicit the release of metabolic mediators with vasodilatory effects, such as increased partial pressure of carbon dioxide potassium, adenosine, and lactate concentrations (Sarelius & Pohl, 2010). Lactate is considered a residual product of metabolism, but today is recognized as a multifaceted molecule, serving as fuel for different cells, with neuroprotective action, hormonal action, anti-aging, metaboreceptor stimulant and vasodilator (Devereux, Coleman, Wiles, & Swaine, 2012;Proia, Di Liegro, Schiera, Fricano, & Di Liegro, 2016). It can act as an important dilator of vascular smooth muscle, however the increase in blood lactate concentration does not seem to influence muscle blood flow, making it difficult to understand its role as a mediator of hyperemia in exercise, as well as its global effect on hemodynamics after a period of training (Lott et al., 2001).
In four weeks of isometric training on isokinetic dynamometer in normotensive young men, significant changes in blood lactate concentration accumulation and reductions in resting systolic BP were observed, in addition to an inverse correlation between lactate concentration and BP changes (Devereux et al., 2012). However, we still do not know whether IHG training has the same effect in hypertensive or prehypertensive elderly people with or without Development, v. 9, n. 9, e655997433, 2020 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v9i9.7433 6 medication, as well as the isolated and combined role of different muscle contraction metabolites, such as NO-, redox balance equivalents and lactate. The hypothesis of this study was that IHG training would contribute for reducing BP and that both lactate concentration and redox balance changes could be associated with this phenomenon.

Methods
The study methodology characterized of an exploratory research, semi experimental study. In relashoship of a nature this study caracterized of a quantitative. That Characterized conformed Pereira et al. (2018).

Participants
Thirty-three elderly people (aged 75.3 ± 1.3 years; Table 1) sedentary with less than 2 h·wk-1 of physical activity volunteered to participate in this study. Everybody lived in a longstay care institution for elderly people and had a controlled routine with predetermined schedules for sleeping, waking, and feeding. Participants were classified in prehypertensive and hypertensive individuals according to the 7th Brazilian Guideline of Arterial Hypertension (Malachias, 2016).

Experimental design
The participants were allocated based on a nonrandomized controlled trial: (a) 11 hypertensive individuals in non-exercise control group (CG); (b) 10 prehypertensive group (PHG); and (c) 12 elderly people in the hypertensive group (HG). The hypertensive individuals (CG and HG) received antihypertensive medication and had been hypertensive for 8.7 ± 1.3 years. The PHG group did not use any type of drug. PHG and HG performed the IHG training, while non-exercise CG maintained its usual activities for the same period.
Individuals with a systolic BP (SBP) > 160 mm Hg or diastolic BP (DBP) > 100 mm Hg, osteoarticular problems, compromised auditory acuity, diabetes, obesity, cardiac arrhythmias, end-organ injury, peripheral arterial disease, myocardial infarction, stroke, coronary heart disease, heart failure, or a recent history of smoking, drug, or alcohol abuse were excluded.
The study consisted of four phases: (i) physical assessment, application of a medical questionnaire, cardiovascular and biochemical measurements; (ii) familiarization with Research, Society and Development, v. 9, n. 9, e655997433, 2020 (CC BY 4.

Outcome measures
Subjects did not perform any physical activity for at least 24 h before the evaluations and avoided caffeine. All data were collected in a long-stay care institution for elderly people, located in Brasilia, Brazil.

Anthropometry
Body mass and height were measured before and after training period, according to the World Health Organization guidelines (Chalmers et al., 1999). Body composition was determined using an adipometer Lange skinfold caliper (Beta Technology, Santa Cruz, CA, USA) using the Durnin and Womersley skinfolds protocol (Durnin & Womersley, 1974) and the Siri equation (Siri, 1961).

Biochemical dosages
A volume of 5 mL venous blood (EDTA, BD Vacutainer®, IL, USA) sample was collected on resting pre and 48 hours after the last training session, at 06:00 AM, with 12 hours of fasting. After collection, venous blood samples were immediately processed in a refrigerated centrifuge to obtain blood plasma (4ºC for 15 minutes at 1500 rpm). All the biochemical analyses were performed at Molecular Biology of Exercise Laboratory of Catholic University of Brasilia.

Resting Blood lactate
The concentration of lactate (mmol · L-1) in the blood was determined in 25μl of capillary blood collected from the distal part of the thumb. Samples were deposited in a plastic tube (Eppendorf®, Berlin, Germany) with 50μl of 1% sodium fluoride, and stored at -80°C for further analysis on an electrochemical biochemical analyzer (YSI 2700 Select, Yellow Springs, OH, USA). The intraassay coefficient of variation was ≤ 10%.

Antioxidant parameter
Total antioxidant capacity (mM) was measured with a Trolox-equivalent assay kit (QuantiChrom® BioAssay Systems, CA, USA). The principle of the antioxidant assay is formation of a ferryl myoglobin radical from metmyoglobin and hydrogen peroxide, which oxides the ABTS (2,2'-azino-bis (3-ethilbenzthiazoline-6-sulfonic acid) to produce a radical cation, ABTS•+, a soluble chromogen that is green in color and was determined spectrophotometrically at 405 nm. The intraassay coefficient of variation was 20%.

Lipid peroxidation
The calorimetrical method for the test of lipoperoxidation by thiobarbituric acid reactive substances (TBARS; nmol.mL-1) was based in the reaction between the trichloroacetic acid (TCA) 17.5%, thiobarbituric acid (TBA) 0.6% and the plasma sample that form a rosy chromogen compound. After homogenization, the samples were kept in a water bath for 20 minutes at 95°C. The reaction was interrupted with the immersion of the microtubes in ice and the addition of TCA 70%, and another incubation for 20 minutes at Research, Society and Development, v. 9, n. 9, e655997433, 2020 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v9i9.7433 room temperature. After centrifugation (3000 rpm for 15 minutes), the supernatant was removed and put in new microtubes and read by spectrophotometry at 534 nm. The concentration of lipid peroxidation products was calculated using the molar extinction coefficient equivalent for malondialdehyde (MDA-equivalent = 0,156 nmol · mL-1). The intraassay coefficient of variation was 7%.

Nitric oxide
NO-bioavailability was measured by plasma nitrite (NO2-; μM) according the description of the Griess Reagent System Kit (Promega Corporation, Madison, USA). One means to investigate NO-formation is to measure NO2-, which is one of two primary, stable and nonvolatile breakdown products of NO-. This assay relies on a diazotization reaction that was originally described by Griess (Griess, 1879). The absorbance was measured in a plate reader with 490 nm filter. The intraassay coefficient of variation was ≤ 10%.

Maximal voluntary isometric contraction
Participants completed 2 weeks of familiarization on handgrip exercise prior to the MVIC. A hydraulic handgrip dynamometer Jamar™® (Sammons Preston, IL, USA) was used to determine the isometric strength; three trials were performed on each hand, and the highest result was recorded (Crosby & Wehbé, 1994;Frederiksen et al., 2006). The positioning of the volunteers while performing the test followed the recommendation of the American Society of Hands Therapists (Fess, 1981). The MVIC test was held before, during (load adjustment every two weeks) and after the training period.

Isometric handgrip training
IHG training was performed during an 8-week period, three times a week on nonconsecutive days. The sessions were held in the afternoon between 02:00 and 05:00 PM.
The volunteers underwent 24 sessions of IHG (Jamar™® Sammons Preston, IL, USA), completing a total of 8 sets of 1-minute contractions at 30% MVIC and 1-minute rest-pause, with 4 sets in each hand, alternately, based on the program described in a previous study (Souza et al., 2018). Volunteers were also instructed to avoid the Valsalva maneuver during the isometric exercise, following ACSM guidelines (Pescatello et al., 2004). The sessions Development, v. 9, n. 9, e655997433, 2020 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v9i9.7433 were supervised by an exercise physiologist and four students of physical education that instructing the volunteers during the exercise.

Statistical analysis
The alpha adopted was 0.05 and the values were expressed as mean and standard error of the mean. Shapiro-Wilk and Levene tests were used to verify the normality and homoscedasticity of the data, respectively, only glucose, lactate, TBARS and Trolox present normality. To compare the hemodynamic response to eight weeks of IHT, we perform the area under the curve (AUC) from the 1st to the 8th week. After calculated, the AUC was compared using ordinary one-way ANOVA followed by Tuckey's post roc. To perform the multiple comparisons between groups over time (group vs. time) to Gaussian variables, it was used the Two-Way ANOVA, with post hoc Least Significant Difference (LSD). For the non-Gaussian variables, we applied the Kruskal-Wallis test followed by Dunn's multiple comparison. Pearson's correlation was used to verify the associations of the study. We performed a backward elimination stepwise multiple regression to determine if we could identify the sequence of variables that could explain blood pressure decrease (Δ NO, Δ lactate, Δ Trolox/TBARS ratio and Δ MVIC relative to lean mass and Δ body mass), at each step, the variable that is the least significant is removed. This process continues until no

Results
The general characteristics of the elderly people are presented in Table 1. There was no intercurrence during the sessions. All participants joined IHG training, completing an average of 88.8% [variance 75% to 95%] of the exercise sessions (~ 21 sessions) during the 8week period. There was no statistical difference between the groups for the anthropometrics variables (P > 0.05). The mean values of hemodynamic variables (Table 1) found in pre-Research, Society and Development, v. 9, n. 9, e655997433, 2020 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v9i9.7433 training were not statistically different between groups (P > 0.05), however, there was a decrease in PHG and HG group in relation to CG (P < 0.05). Date are presented as mean ± SD. CG, control group; PHG, prehypertensive group; HG, hypertensive group; SBP, systolic blood pressure; DBP, diastolic blood pressure; MAP, mean arterial pressure; HR, heart rate; ACE, angiotensin-converting-enzyme; ARB, type 1 angiotensin II receptor blockers; NA, not applicable; * P < 0.05 CG vs. PHG and/or HG (between group); † P < 0.05 (within group). Source: Authors. Table 2 shows the values of the voluntary isometric contraction of both members of the studied groups. In addition, we can also see the basic values performed in three attempts.
There was also no difference, in pre-training, between right and left handgrip strength (P > 0.05) ( Table 2). Handgrip muscle strength was higher in post-training compared to pre- Development, v. 9, n. 9, e655997433, 2020 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v9i9.7433 training in both trained groups, while the CG (non-exercise) decreased muscle strength after 8 weeks. The workload was adjusted every two weeks and increased significantly over baseline in both groups submitted to the IHG training (P > 0.05). Table 2. Neuromuscular variables of participants.
There was no change in Trolox throughout the study (P> 0.05).  Was applied an analysis of backward elimination stepwise multiple regression with Δ lactate, Δ Trolox / TBARS ratio, Δ MVIC relative (kgf • LM-1) Δ nitric oxide and Δ body mass using Δ SBP as dependent variable. As observed in Table 3, the variables presenting significant associations with the decrease in BP were lactate and muscle strength (P< 0.05).
Research, Society and Development, v. 9, n. 9, e655997433, 2020 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v9i9.7433 Clearly, muscle-derived metabolites are formed during exercise, some of them, usually have potential vasodilatation signaling which contributes to exercise hyperemia (Sarelius & Pohl, 2010). Due to the difficult in quantitate the respective contribution of each metabolite in vasodilatation, some studies debate which substances are involved in the regulation of human blood-flow (Mortensen & Saltin, 2014;Nawab et al., 1984;Saltin, Rådegran, Koskolou, & Roach, 1998). Among a number of vasoactive substances, we highlight lactate as an important mediator of vasodilatation (Nawab et al., 1984). It is suggested that lactate stimulate NO synthase and subsequent activation of guanylyl cyclase which will trigger the opening of ATP-sensitive potassium channels of vasodilation (Hein, Xu, & Kuo, 2006). In this regard, the present study adds information regarding the role of lactate in blood flow stimulated by IHG training, which is the variable that most explain BP decrease in both prehypertensive and hypertensive patients. Moreover, Devereux et al (Devereux et al., 2012), suggests that higher levels of anaerobiosis may promote the training-induced reduction in BP which might be linked to increased lactate.
Isometric muscle contraction induces ischemia-reperfusion in the adjacent region (Gaffney, Sjøgaard, & Saltin, 1990), which promotes an increase in the production of free radicals. It is important to induce intracellular adaptations in the antioxidant defense. This can be observed in the work of Peters et al (Peters et al., 2006) in which they demonstrated that the IHG at 50% of MVIC reducing the production of free radicals after 6 weeks of training.
Similar results were found in the present study, however we applied IHG training at 30% of MVIC, demonstrated that even in lower intensities, IHG training can induce systemic and molecular benefits.
It is well known that IHG training reduces BP in hypertensive subjects (Gaffney et al., 1990;Millar et al., 2008;Millar et al., 2014;Peters et al., 2006;Taylor et al., 2003). However, the mechanisms involved in this decrease are not fully elucidated. In this study it was verified that the IHG training in moderate intensity was able to increase antioxidant defense in older prehypertensive and hypertensive patients. Therefore, these results indicate that the BP reduction after eight weeks IHG was partly mediated by the improvement of redox balance, resulting in the increase of the NO-bioavailability, possibly decreasing the total peripheral resistance. Some limitations of study need to be mentioned: i) The heart rate variability has not been evaluated, and it was not possible to use ambulatory BP measurements; however, BP was measured three time a week on nonconsecutive days (the weekly average was used) during IHG training, keeping the same time and place of the long-term care institution for elderly people; ii) This study was not a randomized controlled trial. Therefore, its external validity is limited to this population "prehypertensive and hypertensive elderly people institutionalized". Furthermore, it was not possible to constitute non-exercise prehypertensive control group (a considerable number of volunteers was excluded at the beginning of the project because they were not able to perform the handgrip test due to osteomyoarticular problems in the hands); iii) The sample size limited the analysis of gender influence, of medications use and race in the magnitude of BP reduction. Finally, considering that hypertension is a chronic disease and needs continuous lifelong treatment, it is important to ratify that the long-term effects (> 10 weeks) as well as the adjustment of the volume vs.
intensity during IHG training about BP control in hypertensive elderly people are still unknown.
This study provided important insights of the possible molecular variables that could be associated to BP control. Furthermore, demonstrating IHG training as relevant clinical application due to its potential benefits in BP control, NO-metabolism and redox balance. In this sense, hospitals or clinics would not need to bought strength training machines or weights that could be expensive and take up a lot of place, just with one handgrip dynamometer, prehypertensive and hypertensive patients would be beneficiated by a simple training protocol, which could be applied in any place and any time.
In summary, we conclude that IHG training may be practical in improving clinical status of prehypertensive and hypertensive patients, by improving BP control, NObioavailability and redox balance. Further studies are required to elucidate the pathways of lactate concentration in blood-flow during exercise.
In suggestion for future researchs we believe that other type of exercise or training models can also be another prospect for new studies. We must emphasize that both dynamic strength, aerobic and combined training have already found quite positive effects in relation to hemodynamic aspects in hypertensive patients (Schroeder et al., 2019;Sabbahi et al.,2016;Ferrari et al.,2017). Based on this assumption, the use of different types of exercises can be somewhat effective in the metabolic parameters such as the lactate level. Another question is also the intensity of the exercise for hypertensive people, which is also already described in the literature as an extremely important factor in the responses of the physical exercise (Boutcher & Boutcher, 2017). Other hemodynamic assessments such as heart rate variability, the use of echocardiography may be relevant aspects for new studies on the influence of the protocol used in our study its perpetuations. However, further studies are needed to assess the effects of physical exercise on hemodynamic and metabolic aspect and pre-hypertensive and hypertensive individuals, especially in terms of blood lactate values.

Conflicts of interest
No potential conflict of interest relevant to this article was reported.

Funding source
The authors declare there were no sources of funding for this research.