Effects of 12 weeks of back-squat training program on jump performances and bone markers in female students

Badiea Sharaif,Nidhal Jebabli,Saber Abdellaoui,Jed Mohamed Tijani,Jihen Khalfoun,Mohanad Omar,Abderraouf Ben Abderrahman

1Physical Education and Sport Sciences Department,College of Education,Sultan Qaboos University,Al-Khodh Muscat 123,Sultanate of Oman. 2Higher Institute of Sport and Physical Education of Ksar-Said,University of Manouba,Ksar-Said 2009,Tunisia. 3High Institute of Sport and Physical Education of Kef,University of Jendouba,UR13JS01,Kef 7100,Tunisia.

Abstract Background: The training program promoted improvements of jump abilities throughout the musculoskeletal system including bone markers.The aim of this study is to examine both the acute and chronic response of bone markers to resistance training program. Methods:Ten female students (age: 18 ± 0.7 years, body mass: 63 ± 3.6 kg; height: 164 ± 5.2 cm)participated in this study. They were recruited for a back‐squat training program for 12 weeks,two days/week.The full‐back squat protocol consisted of 3–5 sets×3–8 repetitions at 45–55% one repetition maximum. Testing sessions included a 5 jump test (5JT), standing long jump(SLJ),drop jump(DJ),and vertical jump(VJ).Results:Substantial improvements in all testing jumps (5JT: ?10%; P = 0.000; ES = 1.72; SLJ: ?7%; P = 0.000; ES = 1.33;DJ: ?11%; P = 0.000; ES = 0.72; VJ: ?20%; P = 0.000; ES = 1.84) were found during post program in comparison to pre‐program results. Moreover, a significant change (P ≤0.05) of bone markers during post‐exercise compared to pre‐exercise either before or after the training program. Only collagen type I carboxy‐terminal peptide (CICP) levels elevated after the training program (pre‐exercise only) compared to former levels. Conclusion: 12 weeks of back‐squat training program resulted greater acute improvements of jump abilities with adaptation in all musculoskeletal system including bone formation.

Keywords:strength training; jump tests; bone biochemical markers; athletes

Background

It is well known that resistance training programs are effective in improving physical performance [1, 2]. Indeed, resistance training is one of the approach of explosive origin,which are aimed at increasing lower‐body power from the accentuation of the vertical component during the extension of ankle, knee, and hip, as different squat exercises (for example, the full squat) [3]. Increased descending cortical drive, increased alpha motor neuron input, preferential motor unit synchronization, increased motor unit firing rates, and decreased activation threshold for type II motor units have been introduced as central and peripheral adaptations to resistance training [4‐6].

Physical training presents as a source of bone renewal to prevent osteoporosis and bone metabolism problems. In this context, Bone biochemical markers become very attractive tools for investigating the response of bone cells to exercise and training program, particularly the individualization of exercise programs to improve bone health[7‐9]. Exercise helps to increase bone mass by regulating the balance between bone formation and resorption, and this leads to a net gain in bone mass. However, exercise interventions that increase bone mass do not always result in detectable changes in serum bone formation or resorption markers from pre‐ to post‐intervention [10, 11]. In addition, collagen is the major protein in the extracellular matrix of the musculoskeletal tissue, but despite its importance to tissue function,knowledge of the physiological regulation of the amount and turnover of collagen in human beings is still poor [12]. Hypothesized that a critical component in the transfer of force from the contractile units of the muscle out to the tendon and subsequent bone is the connective tissue scaffold that surrounds individual muscle fibers,muscle bundles,and the whole muscle[13].Collagen fibers constitute the principal connective tissue of skeletal muscles and its linkages within and between muscle fibers that provide their strength and stability. In addition, type 1 collagen has the most abundant bone protein and dominates by more than 90% of the proteins in osteoid tissue [14]. In the same context, Tartrate‐resistant acid phosphatase(TRAP) is an iron‐containing enzyme common to the bone. It is insured by cells of monohistiocytic lineage including macrophages and dendritic cells(DCs)[15‐17].This enzyme is also known as type 5 acid phosphatase with two distinct bands (5a; 5b). The difference in bands was caused by the presence of sialic acid residues in band 5a,unlike that of band 5b [18].

The question that arises is how bone response can adapt to an exercise program with very low innervations. Recently, it was believed that interstitial fluid flow gives bone cells sensation during mechanical activity [19]. Cells coordinate their responses to mechanical load by communicating via the gap junctions. A gap junction is a group of ion channels (connexons) through which molecules of less than 1,000 molecular weight passes through. One level of signaling involves an increase in the intracellular calcium in mechanically stimulated cells whose wave is thought to be propagated from cell to cell through the gap junctions [20]. Intercellular channel gap junctions are composed of two hexameric oligomers called connexins or hemichannels, with one each contributed by contacting cells [21]. The most ubiquitous connexin responsible for intercellular mechanotransduction is the gap junction protein connexin 43 (Cx43).The process by which the body converts mechanical loading into cellular responses is called mechanotransduction. These cellular responses, in turn, promote structural changes. Since weak bone can become larger and stronger in response to the exercise training load through mechanotransduction, bones adapting to these loads are examples of mechanotransduction [22].

To the best of our knowledge, no previous studies have compared the acute response of bone markers to chronic response with regard to gap junction functional protein connexin 43. Therefore, the overall objective of the present study is to study the effect of a 12‐week periodized back‐squat training program on jump performances,plasma type I procollagen (CICP; a marker for bone formation),enzyme serum band 5 tartrate resistant acid phosphatase (TRAP 5b; a marker of bone resorption) and connexin 43 (gap junction response).It was hypothesized that back‐squat training program would modify the acute bone formation during post‐program compared to pre‐program in these entire enzyme parameters. It was hypothesized that back‐squat training program would modify the acute bone formation during post‐program compared to pre‐program in these entire enzyme parameters.

Material and methods

Subjects

Based on a medium‐sized effect of strength training program on bone re‐modulation, a power analysis was calculated by G＊Power (Version 3.1.9.2,University of Kiel,Kiel,Germany)using the t‐student test.The analysis revealed that a total sample size of N = 10 would be sufficient to find significant and medium‐sized effects of condition(effect size f = 0.6, α = 0.05) with an actual power of 0.97. Ten healthy active female students were randomly selected(age: 18±0.7 years; body mass: 63 ± 3.6 kg; and height was 164 ± 5.2 cm) from the faculty of physical education for girls at Helwan University in Giza,to participate in this study. The participant voluntarily provided written informed consent before participating. All participants beneficed of a medical exam prior to inclusion that revealed no cons‐indication for physical exercise. Participants were asked to follow their normal diet (10 kcal/kg, 55% of which came from carbohydrates,33%from lipids and 12%from proteins,as determined by an experienced nutritionist) during the program.The present study was conducted according to the latest version of the Declaration of Helsinki,and the protocol was fully approved by the Ethics Committee of the faculty of physical education for girls, Helwan University, Giza,Egypt with the declaration of Helsinki and approved by the local ethics committee (Helwan University, Giza, ID 2019‐133/13) before the commencement of the tests.

Training intervention

The study was designed to test the effects of a 12‐week back‐squat program on selected fitness measures in young active female students.Week prior to the intervention, one session familiarized students with all test procedures, including anthropometric measurement and determination of 1‐RM back‐squat for each student. Measurements of 5 jumps test (5JT), Standing long jump (SLJ), drop jump (DJ), and vertical jump(VJ)tests were made 2 days before and after the training program.Subjects did not participate in any exhausting exercise for 24 h before testing. During training program, student trained twice a week (with 72 h rest between sessions) during 12‐week period, using only the back‐squat exercise. Every session started with 15‐min warm‐up consisting of 10‐min continuous jogging at moderate intensity (50% of maximal aerobic velocity (MAV) followed by 5‐min dynamic stretching exercises. Back‐squat exercises were developed at the gym, using a squat rack machine (FT700 Power Cage, Fitness Technology). Training program design is detailed in Table 1.

Table 1 The strength training program over 12 consecutive weeks

1-RM back-squat test

A familiarization protocol was applied prior to the beginning of the study. This protocol consists of the familiarization of 1‐RM back‐squat test. Participants were asked to descend to where the upper thighs were horizontal or lower, followed by an ascent until the hips and knees were fully extended. Participants were asked to control the rate of descent and to raise the bar as quickly as possible, with control,during the ascent.1‐RM was calculated using the following formula of Brzycki[23].

1RM = 100 × load rep/(102.78 – 2.78 × rep)

where:

?load rep:workload value of repetitions performance,expressed in kg;

?rep: number of repetitions performed.

Back-squat training program

The back‐squat training consisted of 3–5 back‐squat sets × 3–8 repetitions at 45–55% 1RM (Table 1) [24]. The concentric phase was performed as fast as possible, and the eccentric phase was executed in a controlled manner (i.e., approximately 2 seconds). The resting period between each set was 3 minutes. Finally, participants were cooled down by running at low intensity and performing static stretching for 10 min.

Testing sessions

5 jump test(5JT).The 5JT consists of 5 successive strides with joined feet position at the start and the end of the jumps. From the starting joined feet position, participant was not allowed to perform backward movements; he had to directly jump to the front with the preferred leg. After the first 4 strides, the participant completes the last stride with joined feet. If the player backs up at the end of the last stride,the test is carried out again.

Standing long jump (SLJ).The standing long jump was used as a power test for the bilateral legs.Arms movements were authorized for support during takeoff movements. The trials were evaluated only when the subjects landed correctly on their feet without folding back.The best of two SLJ trials was used as the best performance[25].

Vertical jump (VJ).With the side preferred on the wall, the vertical jump test or Sargent jump test is essentially consist of measuring the difference between a person’s standing reach (first mark) and the height at which they can jump and touch (second mark) [26]. The vertical jump score is the difference between the two marks (recorded in centimeters).

Drop jump(DJ) test.The participants performed DJ by first jumping from the box (30 cm in high), landing and immediately performing a maximum vertical jump.No specific information was provided on how to land or jump. This test was repeated 3 times. The best of DJT trials was used [27].

Blood analysis

Blood samples at rest were collected under aseptic conditions from the antecubital vein. Post‐exercise blood sampling was performed immediately after the exercise unit. Venous samples for the measurement of plasma parameters were collected in tubes containing EDTA in the morning after fasting, then after resting for at least 30 minutes and immediately after the exercise, it was then centrifuged and kept at –20 ℃until tested. Type I procollagen, TRAP 5b and Connexin 43 levels were determined using ELISA kit technique provided by Teco medical group, Germany [28].

Results

Physical parameters results revealed significant variation in all parameters (P< 0.01) during testing post‐program compared to that obtained during pre‐program(Table 2).

Table 2 Comparison between physical parameters(pre-program related to post-program)

Revealed data results indicated no significant variations in the pre‐program blood levels of tartrate‐resistant acid phosphatase 5b(TRAP b5) and connexin 43 compared to that obtained post program either at rest (Pre‐exercise) or after exercise (post‐exercise) but were significant for CICP(?=22%;P=0.035＊; effect size=1.60)only in pre‐exercise after the program (Table 3).

Table 3 Comparison between blood levels of TRAP b5, CICP and Connexin 43 pre-program to post-program

Reverse to the obtained results of pre‐to post‐program, pre‐exercise compared to post‐exercise either before or after the program were significant in all the three investigated bone parameters as presented in Table 4.

Table 4 Comparison between blood levels of TRAP b5,CICP and connexin 43 pre-exercise to post-exercise

Discussion

The main purpose of this study was to analyze the effect of a 12‐week periodized back‐squat training program on jump performances,plasma type I procollagen, TRAP 5b and connexin 43. In agreement with our research hypothesis, the main results of this study suggest that back‐squat training program developed significantly jump performances compared to that obtained during pre‐program. Also,the present study showed a significant change (P≤0.05) of bone markers during post‐exercise compared to pre‐exercise either before or after the training program.

The present results indicate that 12 weeks of back‐squat training program had a beneficial impact (P= 0.000) on 5JT, SLJ, DJ, and VJ performances (↑?: 7‐20%; ES: 0.72–1.84). In other words, the use of medium loads (45–55% of 1‐RM) with rapid actions to induce maximum neural adaptation can improve significantly jump performances. In agreement with the present results, previous investigations have reported typically greater training effects of jump performance after a resistance training program [29‐31].

Our results are an agreement with previous studies like Burgomaster et al.who proved that strength or power training produces marked increases in muscular strength which could be attributed to a range of neurological and morphological adaptations and it was one of the most widely practised forms of physical activity, which is used to increase musculoskeletal health and alter body aesthetics, enhance athletic performance[32].Moreover,TRAP b5 was evaluated for bone resorption, CICP for bone deposition and connexin 43 for inter‐bone cellular coordination in response to the training program. There were no significant differences observed at rest (pre‐exercise) or after exercise (post‐exercise) in all the results obtained except for CICP(Table 3) compared to themselves, which was found to be elevated at rest after the training program (?= 22%;P= 0.035＊; effect size =1.60). In addition, bone is continually undergoing processes of reinforcement and resorption (bone functional adaptation) until the skeletal design meets the loading requirements. The cellular and molecular studies indicated that bone cells, i.e. osteoblasts and osteocytes, are mechanosensitive cells [33].

In accordance with the present study, Rocha et al. [34] revealed that 2 to 4 months of training period including marching, running,jumping, and battle drills improve significantly TRAP b5 in both men and women.However,no significant change of TRAP b5 was observed after 60 min of cycling ergometer at 75% to 110% of maximal aerobic velocity [35]. The differences between the results of the present study and those mentioned above may be due to the change in the intensity and volume of training program. Previous studies like Lanyon [36]observed that osteocytes appeared to reflect the strain signals and distribute them throughout the whole bone to regulate bone(re)modeling.Recently it was shown that deletion of osteocytes results in bone loss and bone with deleted osteocytes does not respond to load[36]. These studies show that osteocytes are necessary to maintain bone mass in response to normal load,but in the absence of load,they send signals of resorption [37]. Hence, as revealed in the present study,the net result of training program leads to bone deposition with elevated CICP as bone formation marker. We hypothesized that exercise would modify the acute bone response that resulted in post‐exercise compared to pre‐exercise in all the investigated parameters(Table 4).These results seem to be related to the timing of the measurement of bone turnover markers relative to the last exercise bout or the diurnal variation of bone resorption markers rather than the training program effect [37].

In the literature, Rogers et al. studied the acute response of bone formation and resorption markers including tartrate‐resistant acid phosphatase 5b (TRAP 5b), COOH terminal telopeptide of type I collagen (CTX), to a single bout of resistance exercise or polymetric exercise [10]. In general, they observed cumulative decrease in TRAP 5b during the 2 h exercise while CTX remained unchanged. Their results suggested that the timing of the measurement is important for detecting exercise‐associated changes in bone turnover markers,as the markers returned to pre‐exercise values within 2 h of exercise. Type I collagen synthesis response is accelerated by prolonged strenuous exercise, peak after 3 days and lasting for 5 days to re‐return to basic levels[38]. Moreover, Virtanen et al. [39] observed that a single bout of heavy concentric exercise causes protein leakage from muscles and probably from the collagen‐synthesizing cells of the connective tissue,which may be accompanied by an initial decrease and a subsequent increase in type I collagen production. The activation of type I collagen production seems to depend on the strain and damage of the musculoskeletal system [40]. Plasma connexin 43 was the first to be estimated as a marker for gap junction in response to the training program. The results were not significant post program compared to pre‐program. These results may be explained under the hypothesis that it affects bone mass rather than releasing the lymphatic system.Connexin 43 junctional conductance is activated at 70‐85 mV which causes junctional currents, resulting in a steady‐state conductance which is a fraction of the instantaneous conductance [41]. Connexin enhances fluid flow through the canalicular system of the osteocytes to amplify the strain signals that induces increase in bone mass.Virtanen et al.revealed that loss of connexin 43 expression resulted in poor junctional competence and loss of the ability to transmit a calcium signal to a neighboring cell. Transfection of junctionally incompetent cells with connexin 43 DNA codon restoring intercellular communication. Therefore, gap junctions represent one mechanism cells that are used for regulating a response to mechanical and chemical signals [40].

Some limitations of this study should be acknowledged. Indeed, the results of this study are limited by the small number of populations analyzed.Also,it is important to evaluate neuromuscular performance such as flexibility, muscle strength in relation to the variation of bone markers.

Practical applications

This study suggests that 12‐week periodized back‐squat training program was efficient in attenuating the raise in bone remodeling and has an effect on jump performances. This training program can prevent old bone tissue from accumulating in the skeleton and helps more in the formation of new bone tissue. Our results may be relevant in the practical application of this training. Strength training program can be used easily by coach and personal trainer for female active and sedentary. In relation avec jump tests, a good familiarization of the exercises before the training program will be very necessary to avoid muscle and bone injuries.

Conclusion

Future research should clarify the evaluation of calciotropic hormone(vitamin D and calcitonin) necessary to bone remodeling.