Muscular Strength Adaptations and Hormonal Responses After Two Different Multiple-Set Protocols of Resistance Training in Postmenopausal Women

Abstract

Nunes, PRP, Barcelos, LC, Oliveira, AA, Furlanetto, R, Martins, FM, Resende, EAMR, and Orsatti, FL. Muscular strength adaptations and hormonal responses after two different multiple-set protocols of resistance training in postmenopausal women. J Strength Cond Res 33(5): 1276–1285, 2019—We studied the effects of 2 different resistance training (RT) multiple-set protocols (3 and 6 sets) on muscle strength and basal hormones concentrations in postmenopausal women (PW). Postmenopausal women were randomly allocated into 3 groups: control (CT, n = 12), low RT volume (LV = 3 sets for each exercise, n = 10), and high RT volume (HV = 6 sets for each exercise, n = 12). The LV and HV groups performed 8 exercises of a total body RT protocol 3 times a week, at 70% of 1 repetition maximum (1RM) for 16 weeks. Muscle strength and basal hormone concentrations were measured before and after RT. Our findings show that 3 sets or 6 sets at 70% of 1RM protocol increased muscular strength similarly after 16 weeks (sum of all exercises, LV: 37.7% and HV: 34.1% vs. CT: 2.1%, p < 0.001). Moreover, the RT volume does not affect basal levels of testosterone (TT) (LV: 0.02%, HV: −0.12%, and CT: 0.006%, p = 0.233), cortisol (C) (LV: 72.4%, HV: 36.8%, and CT: 16.8%, p = 0.892), insulin-like growth factor-1 (LV: 6.7%, HV: 7.3%, and CT: 4.1%, p = 0.802), dehydroepiandrosterone sulfate (LV: 0.1%, HV: −4.5%, and CT: −6.7%, p = 0.885), and TT:C ratio (LV: −0.9%, HV: −1.6%, and CT: −0.4%, p = 0.429). Our results suggest that 3 sets and 6 sets at 70% of 1RM seem to promote similar muscle strength gain. Thus, 3-set RT is a time efficient protocol for strength gain after 16 weeks in PW.

Introduction

In women, a decline in estrogen production from menopause induces a phase of rapid decrease in muscle strength (^3,42,50). Low muscle strength is an important determinant of disability and low quality of life and is also associated with an increased risk of falls, osteoporotic fractures, and mortality in older women (^{3,14,32,36,50}). Resistance training (RT) is an effective method to improve muscle mass and strength, and it is also often prescribed for enhancing physical capacity (i.e., defined by the Centers for Disease Control and Prevention as difficulty in performing physical tasks) and preventing falls, osteoporosis, and mortality in postmenopausal women (PW) (^1–3,5,51).

Muscle strength gains by RT depend on numerous acute training variables such as load (percentage of 1 repetition maximum [1RM]) and volume (amount of exercise; i.e., sets × repetitions × load) (^1–3,5,51). Although the role of moderate-to-heavy loads (>70% of 1RM) in muscle strength gains are well established in older adults (^1,2,5,51), little is known about the effects of RT volume on muscle strength gains. The meta-regression study of Borde et al. (⁵) compared RT protocols (sets ranging from 1 to 5 sets) and observed that 2–3 sets per exercise result in the largest improvement in muscle strength; nevertheless, the author pointed out that there is a paucity of data from high-quality randomized clinical trials (RCTs) concerning the effects of RT volume on muscle strength, especially in the elderly. To the best of our knowledge, only 5 studies have investigated the effects of RT volume on muscle strength gains in older adults (^{10,17,43,44,47}). However, no RCT studies investigating the RT volume effects on muscle strength in older people have used more than 3 sets. These studies have shown that low-volume RT, even as low as 1 set, increases muscle strength in older people (^{10,17,43,44,47}). However, 3 of these studies have confirmed additional gains in muscle strength (of approximately 40%) with a 3-set RT protocol in comparison to single-set protocols (^17,43,47). Consequently, the RT protocol with moderate-to-heavy loads and 3-set volume has been recommended to counteract sarcopenia (^1,2,5,51).

Resistance exercise (multiple sets and >30% 1RM) is a potent stimulator of muscle protein synthesis (MPS) (^8,9,29). Consequently, repeated resistance exercise bouts result in muscle hypertrophy due to cumulative periods of high rates of MPS that exceed the rate of muscle protein breakdown (MPB) (periods of positive net protein balance) (^4,12,13). Older adults, particularly PW, have lower muscle strength gains and hypertrophy than young adults after RT owing, at least partially, to reduced MPS responses (anabolic blunting) (^{4,16,25,26,29}). Interestingly, recent evidence has shown that by increasing the RT (multiple sets, >40% 1RM, >6 repetitions, with 3 minutes rest between sets) volume from 3 to 6 sets enhances postexercise MPS in older people, reaching MPS values similar to young adults (²⁸). Therefore, it would seem reasonable to assume that higher volume RT (6 sets) as opposed to the recommended volume (3 sets) would promote an optimal response for hypertrophy gains and, consequently, muscle strength in PW.

Anabolic hormones (e.g., testosterone [TT] and dehydroepiandrosterone [DHEA]) and growth factors (e.g., insulin-like growth factor [IGF]-1) promote an increase in MPS and also activation, proliferation, and differentiation of myogenic satellite cells maintaining muscle tropism (⁴⁹). This is important because in older adults, particularly PW, basal anabolic hormones and growth factors are diminished, and it is thought that this is the cause of sarcopenia (^19,23,42). Suppression of basal total and free TT results in decreased MPS, muscle hypertrophy, and strength (³¹), affecting muscle adaptation after RT (^7,30,35). Interestingly, some studies have shown that basal circulating hormone levels may play a role in muscle strength gain and hypertrophy during RT in older women (^{20,21,38–40}). For instance, Häkkinen et al. found a positive correlation between basal total and free TT levels and changes in maximal strength (²¹) and muscle hypertrophy (²⁰) after 21–24 weeks of RT (load: 50–80% 1RM, volume: 4–6 sets, rest: 2 minutes between sets) in older women. Furthermore, Orsatti et al. also observed a positive association between a basal IGF-1, total TT, and DHEA levels with changes in muscle mass after 16 weeks of RT (load: 60–80% 1RM, volume: 3 sets, rest: 1.5–2 minutes) in PW (^38–40). Thus, low levels of basal anabolic hormones, especially in PW, may limit muscle mass and strength gains from RT. Therefore, as there is a relationship between basal anabolic hormones and muscle mass and strength gains after RT, it would seem reasonable to assume that these basal circulating hormone (TT, IGF-1, and DHEA) levels are confounding variables of RT muscle adaptation after RT in PW. However, no RCT studies investigating the RT volume effects on muscle strength in older people have controlled the circulating basal hormone (TT, IGF-1, and DHEA) levels as a confounding variable of RT adaptation (^{10,17,43,44,47}).

The dose-response benefits of exercise can be estimated based on RCT studies. However, no RCT study has used more than 3 sets in older adults (^{10,17,43,44,47}). Considering this, the efficacy of RT protocol should be measured by the protocol's ability to increase muscle strength, comparing the effect of the experimental protocol (i.e., high volume [HV]: 6 sets) with a gold standard protocol (i.e., low volume [LV]: 3 sets) (^1,2). To confirm if higher volume RT would lead to superior gains in muscle strength in PW, we examined the effects of 2 different RT multiple-set protocols (3 and 6 sets) at the same intensity (70% of 1RM) on muscle strength gains and basal hormone concentrations (as a confounding variable of RT adaptation) in PW. Acknowledging that the RT volume is an important stimulus for promoting muscular adaptation in PW, we hypothesized that greater gains in muscle strength could be detected when the RT is performed using 6 sets.

Methods

Experimental Approach to the Problem

This study was conducted to evaluate the effect of RT volume on muscle mass and strength in PW. A randomized, controlled study was performed over 16 weeks. Muscle strength (1RM test), lean body mass (LBM), and circulating basal hormone measures were assessed at the baseline and at the end of the RT intervention. The sample consisted of 34 PW divided into 3 groups: control (CT, n = 12), LV (n = 10), and HV (n = 12). The LV group performed a total body RT protocol (8 exercises) with 3 sets of 8–12 repetitions at 70% 1RM with 1.5 minutes rest interval between sets and exercises, 3 times a week. The HV group performed the same RT as the LV group, except for the number of sets that were 6. The total training volume was calculated by multiplying the load by sets by repetitions. The CT group did not participate in the RT routine; they only did stretching exercises twice a week. After the RT period, the assessments were performed 72 hours after the last session of training to avoid workout side effects.

Subjects

All the volunteers were sedentary women (age range = 49–79 years old) whose amenorrhea had occurred at least 12 months before the study and whose basal follicle-stimulating hormone (FSH) and basal estradiol (E₂) values were >40 mIU·ml⁻¹ and ≤54.7 pg·ml⁻¹, respectively, to determine menopause status. The inclusion criteria consisted of no hormone therapy or phytoestrogens; controlled blood pressure and glycemia; absence of myopathies, arthropathies, and neuropathies; absence of muscle, thromboembolic, and gastrointestinal disorders; absence of cardiovascular and infection diseases; and nondrinker (no alcohol intake whatsoever in their diet) and nonsmoker. Before the study, basal thyroid-stimulating hormone (TSH) and basal free thyroxine (T4) levels were measured to exclude thyroid dysfunctions that could interfere with the symptoms. A CONSORT diagram is shown in Figure 1. Thus, the 34 volunteers who participated in the study demonstrated E₂ and FSH values within the normal range for PW (Table 1). Twenty-three medication users were identified. Five used glucose-lowering drugs (CT = 2, LV = 2, and HV = 1), 7 used lipid-lowering drugs (CT = 4, LV = 2, and HV = 1), 4 used anti-inflammatory drugs (CT = 1, LV = 2, and HV = 1), and 19 used blood pressure-lowering drugs (CT = 7, LV = 6, and HV = 6).

Figure 1.: Participant flow diagram.

Table 1.: Baseline characteristics of postmenopausal women in the control, low-volume, and high-volume resistance training groups.*†

All PW were clear on the objectives and procedures of the study and gave us their written informed consent. The study was approved by the Federal University of Triangulo Mineiro (UFTM) Review Board for the Use of Human Subjects (local Ethics Committee) and was written in accordance with the standards set by the Declaration of Helsinki.

Procedures

Nutritional Assessments

All women underwent a 3-day food record (2 days in the middle of week and 1 at the weekend) (⁵²). Energy and macronutrients (carbohydrates, proteins, and fats) were quantified. Macronutrients data were corrected for body mass to reduce the interindividual differences. Data were calculated by a nutritionist and the software “Dietpro” 5i version was used. The women were advised to maintain the initial nutritional habits until the end of the study so as not to affect the outcome measures.

Anthropometric Assessments and Maximum Strength Assessment

Concerning the anthropometric assessments, the body mass and height were measured using a digital scale (Lider, Araçatuba, Brazil) and a stadiometer fixed to the scale, respectively. Body mass index was classified according to the system used by the World Health Organization (³⁷). The skinfolds were evaluated using an adipometer (Lange, São Paulo, Brazil) on the right side of the body and was performed at 4 sites (biceps, triceps, subscapular, and suprailiac) by an experienced examiner. Three measurements were taken on each skinfold, and the mean value was taken. The body density was determined according to the equation proposed by Durnin and Womersley (1974) (¹⁵), and the body fat percentage was determined using the equation proposed by Ortiz et al. (⁴¹). Both equations were selected specifically for the population of this study. Lean body mass was estimated from the difference between body mass minus fat mass.

In the maximum strength test (1RM), the maximum load that an individual could lift in a given exercise, involving muscle groups either using free weights or muscle-building apparatus, was quantified. Before the 1RM test, the women attended a 2-week familiarization period. In the first week of the familiarization period, the women performed 3 sessions on alternate days with low loads to learn the exercise techniques and to familiarize themselves with the free weights and muscle-building machine (Buick Fitness, Rio de Janeiro, Brazil) exercises. In the second week of familiarization, the women performed 3 sessions on alternate days to familiarize themselves with the 1RM test technique for each exercise in the following order: 45° half squat (Smith machine), leg curl, leg extension, bench press, rowing machine, pull down, barbell curls, and triceps pulley. In this test, a subjective low load (estimated loads of 40–60% of the 1RM) was determined for the warm-up, and 5–10 repetitions were performed. After the warm-up, the women were allowed to rest for 1 minute. Afterward, 3–5 repetitions were performed, and the subjective load was increased progressively between 60 and 80% of 1RM. After doing this exercise, the women rested for 3 minutes. Then, the load was increased considerably as close as possible to the individual's maximum capacity, and the volunteers attempted to perform the movement. When the load was overestimated or underestimated, the volunteers rested 3–5 minutes before they attempted it again with a lower or higher load, respectively. This procedure was followed to find the equivalent load of 1RM, which ranged between 3 and 5 attempts. The load that was adopted as the maximum load was the one used for the last part of the exercise that was performed with no more than 1 repetition by the volunteer (⁴⁰). During the test, the individuals were advised to avoid respiratory apnea.

Resistance Training Protocol

The RT protocols follow the recommendations of the American College of Sports Medicine Guidelines for muscle hypertrophy and strength (²). No exercise other than RT was allowed, and there was no muscular or joint damage caused to the volunteers. The total body RT protocol consisted of dynamic exercises for the trunk and upper and lower limbs. A 3-day-a-week (no consecutive days) regime of the RT protocol was performed over 16 weeks. All workouts were supervised by a qualified professional. A warm-up session (1 set of 15 repetitions) with 40% of 1RM was preformed in each exercise before each RT session. At the end of the RT sessions, stretching exercises were preformed so that participants could cool down. Both trained groups performed the RT protocol with 70% 1RM. The LV group performed 3 sets (RT length time ∼45 minutes) and the HV 6 sets (RT length ∼90 minutes) of 8–12 repetitions. The HV group started with 3 sets and increased 1 set per week until they did 6 sets for all exercises (week 4). A resting period of 1.5 minutes was established between the sets and exercises. During the workout, the participants were advised to perform eccentric actions in 1 second and concentric actions in 1 second. During the training period, in the eighth week, the load was adjusted according to the 1RM test to keep the relative load of 70% of 1RM and to ensure a progressive overload.

Blood Samples

Blood samples (16 ml) were collected by a qualified professional in the morning between 7:30 am and 9:00 am after overnight fasting (10–12 hours). All volunteers were advised to maintain hydration a day before and on the day of the blood collection. The blood samples were collected by venous puncture in a vacuum-sealed system (Vacutainer; Becton Dickinson, Franklin Lakes, NJ, USA) into a dry tube using a gel separator. The sample was centrifuged for 10 minutes (1,108g), and samples were separated and stocked for 6 months (−20° C) for future analysis. At the end of the study, the last samples were collected after 72 hours after the last RT session.

The basal hormones TSH, T4, E₂, FSH, and DHEA-sulfate (DHEA-S) were measured using Cobas 6000 equipment with Roche (USA) kits for each specific hormone; the calibrators were run once per reagent lot using fresh reagent with no more than 24 hours because the reagent kit was registered on the equipment. The intra assay and inter assay variances for TSH, T4, E₂, FSH, and DHEA-S were 3.2–7.2, 1.6–6.3, 1.3–7.0, 2.5–4.5, and 2.3–3.2%, respectively. The other basal hormones cortisol, IGF-1, and total TT were measured using the Readwell Touch equipment (Robonik, Navi Mumbai, India) with DRG kits (USA). The intra assay and inter assay variances for cortisol, IGF-1, and total TT were 3.2–8.1, 4.7–7.7, and 3.2–9.9%, respectively. All samples were run with a single detection.

Statistical Analyses

The data are presented by mean values and confidence interval (CI) of 95%. One-way analysis of variance (ANOVA) was used for comparisons between groups at baseline characteristic variables. Repeated measures ANOVA was used to determine the interaction of time (pre and post) by group (CT, LV, and HV) effect. ES were measured by partial eta-squared () (¹¹). The observed power was also calculated for this study. The statistical significance was considered at p ≤ 0.05.

Results

The initial characteristics of all volunteers are shown in Table 1. In all the PW, 23.5, 35.3, and 41.2% were classified as eutrophic, overweight, and obese, respectively. There were no differences between the groups considering food intake (Table 2), strength (Table 3), LBM (CT = 45.6 [CI 95%: 40.9–50.2] kg; LV = 40.5 [CI 95%: 37.3–43.6] kg; HV = 41.2 [CI 95%: 37.2–45.2] kg; p ANOVA = 0.121), and basal hormones (Table 4) at the baseline.

Table 2.: Baseline and post values and magnitude of change (Δ%) for dietary intake after the 16-week resistance training period.*†

Table 3.: Baseline and post values, magnitude of change (Δ%), and accompanying effect size for strength performance after the 16-week resistance training period.*†

Table 4.: Baseline and post values and magnitude of change (Δ%) for hormones after the 16-week resistance training period.*†

The HV group showed a higher total training volume (∼2-fold higher) when compared with LV in weeks 4, 8, and 16. The LV group showed increased total training volume (1.2-fold higher) in weeks 8 and 16 when compared with week 1. The HV group showed an increase in the total training volume (∼2-fold higher) in weeks 4, 8, and 16 when compared with week 1. Furthermore, the HV group increased the total training volume (1.1-fold higher) in weeks 8 and 16 when compared with week 4 (Figure 2).

Figure 2.: Comparison of resistance training volume during 16 weeks. CT = control group, LV = low volume group, HV = high volume group. *Significantly different from week 1, p ≤ 0.05; #Significantly different from week 4, p ≤ 0.05.

After 16 weeks of training, a significant interaction of time by group effect was observed for 1RM performance and for LBM. The HV and LV groups increased 1RM performance in all exercises when compared with the CT group having significant effects. However, there were no differences in strength gains between the HV and LV groups, except for the barbell curl that increased only in LV (Table 3). The HV and LV groups increased the LBM when compared with the CT group (CT = 0.8 [CI 95%: −0.1 to 1.8] kg; LV = 2.2 [CI 95%: 1.7–2.7] kg; HV = 2.4 [CI 95%: 1.2–3.6] kg p ANOVA = 0.037). There were no significant time interactions by the group for food intake and hormonal responses (Tables 2 and 4).

Discussion

We studied the effects of 2 different RT multiple-set protocols (3 and 6 sets) on muscle strength gains and hormonal responses in PW. The main finding of our study was that the 6-set RT protocol did not promote additional effects for muscle strength gains and hormonal responses in comparison with the 3-set RT protocol after 16 weeks for PW.

Previous research was conducted to determine the influence of RT volume on muscle strength gains in older adults (^{1,2,5,17,29,43,47}). Most studies have pointed out that a 3-set protocol is better than single-set protocols (^{1,2,5,17,43,47}), establishing a relationship between dose (volume) and response (muscle strength gains) (²⁸). However, to the best of our knowledge, the effect of high-volume RT on muscle strength gain in older people has not been shown because no RCT studies have used more than 3 sets. Thus, our RCT study is the first to show the effects of 6-set volume RT and 3 sets on muscle strength in PW. We demonstrated that both RT protocols (3 and 6 sets) similarly improved the muscle strength gains when compared with the CT group. Although we believe that the training volume is important for muscle strength gains, our study suggests that adding sets, as well as 3 sets at 70% of 1RM promotes no additional effects for muscle strength gains in PW. In accordance with our study, the meta-regression study by Borde et al. (⁵), comprising 25 studies conducted with older people (using a number of sets ranging from 1 to 5 sets), observed that 2–3 sets per exercise result in the largest improvement in muscle strength.

Muscle hypertrophy (which may be considered as a contributor of muscle strength gains) emerges as a result of high rates of MPS that exceed the rate of MPB after resistance exercise (^{1,6,9,12,32–34}). Kumar et al. (²⁸) showed that increasing the RT volume from 3 to 6 sets increases postexercise MPS in older men, reaching MPS values similar to that in young men. As older adults have reduced MPS response (anabolic blunting) (^16,25,26,29), it would seem reasonable to assume that strategies to increase MPS (such as higher RT volume) may enhance muscle mass and strength gains. However, we did not find any differences in LBM and muscle strength gains between 3 and 6 sets. Recently, Damas et al. (¹²) showed that MPS response in an initial RT program is not meant to support muscle hypertrophy, coinciding with the greatest muscle damage. This suggests that the high MPS in a 6-set protocol when compared with a 3-set protocol in a study performed by Kumar et al. (²⁸) may be due to high-volume RT-derived muscle damage. Indeed, Roth et al. (⁴⁸) showed that older women have higher levels (∼2 fold) of muscle damage after 9 weeks of high-volume RT (5 sets) than young women. Moreover, Krieger (²⁷) analyzed 8 studies in a meta-analysis with training sessions ranging from 6 to 24 weeks in young and middle-aged adults (19–45 years) and demonstrated that muscle hypertrophy was no different between 2 and 3 sets compared with 4–6 sets. Although anthropometric analysis may be a limitation of our study to estimate LBM, the relative hypertrophy corresponded to approximately 5.5% (LV = 5.5 [CI 95%: 4.1–6.9] %; HV = 5.7 [CI 95%: 3.0–8.3] %), which was similar to the relative hypertrophy of 4.0–5.5% observed in other studies performed with elderly people (⁴⁶) and PW (²²), which used magnetic resonance imaging and ultrasound, supporting our results.

We demonstrated that LBM and strength gains were similar between the different volumes (6 sets vs. 3 sets) after 16 weeks in untrained PW. Our results are consistent with those observed in short-term RT (<12 weeks), suggesting that a number of sets is not the primary variable responsible for increases in LBM and strength during the early phase of RT in older adults (^5,43). However, a few studies have suggested that trained PW need greater volume stimuli than untrained subjects to achieve further gains (^24,43). Radaelli et al. (⁴³) reported that 3-set RT caused more muscle hypertrophy (∼5%) and strength gains (∼20%) when compared with single-set RT solely after 20 weeks of training. This suggests that the effect of volume may be observed solely after a minimum amount of weeks (i.e., 20 weeks). Thus, the lack of differences between RT volumes (3 sets vs. 6 sets) in our study may be due to the short-term period of RT (16 weeks). However, there is a lack of data on high-quality RCT studies concerning the effects of training volume in long-term RT on muscle mass and strength gain in elderly people. Thus, future research is needed to address this issue.

In older women, muscle mass and strength gain after RT have been associated with nutritional patterns (i.e., protein intake) (²²), basal TT, DHEA-S, and IGF-1 levels (^21,38–40). Thus, low levels of protein intake and basal anabolic hormones, especially in PW, may limit muscle mass and strength gain after RT. However, in this context, no RCT studies investigating the RT volume effects on muscle mass and strength in older people have controlled the nutritional patterns and circulating basal hormone levels as a confounding variable of RT adaptation. In this RCT study, we found no differences between groups in baseline nutritional patterns and basal hormone concentrations and after 16 weeks, which suggests that the muscle mass and strength response observed here were not due to the nutritional patterns and basal hormone concentration differences. Moreover, the possibility of bias due to the measurement error of muscle mass and strength responses was low because of the fact that the assessments were performed by same examiner and the use of a CT group (RCT).

In conclusion, our findings show that 3 sets at 70% of 1RM protocol increase muscular strength similarly to 6 sets at 70% of 1RM protocol after 16 weeks. Moreover, the RT volume does not affect basal levels of TT, cortisol, DHEA-S, IGF-1, and TT:C in PW. Thus, 3 sets at 70% of 1RM seem to promote a RT optimal volume to optimize the muscle strength gains after 16 weeks in PW.

Practical Applications

In the first 16 weeks of training, there was no difference between 3-set and 6-set RT protocols at 70% of 1RM for upper- and lower-body muscle strength gain in PW. Therefore, if the aim is to achieve the optimal strength gain in older women, we recommend 3-set total body RT throughout the first 16 weeks of training. Based on maximal physiological adaptations, a possible volume threshold to elicit greater muscle strength gain seems to be 3 sets, and it is unlikely that RT volume over this threshold will provide further benefits in muscle strength gain in older women in short-term RT. Furthermore, time efficient protocols are more attractive as time to exercise has been a common barrier among people (¹⁸) (duration of the 3-set protocol was 45 minutes and 6-set protocol was 90 minutes). However, PW who need to increase energy expenditure per week would benefit from a progression of RT with a volume ranging from 3 to 6 sets (³⁶) with no detrimental effects on muscle strength gains.