1. Introduction
It is well established that regular physical exercise is of enormous value for the prevention and treatment of numerous chronic diseases. A physically inactive lifestyle is considered a major behavioral risk factor associated with a variety of health conditions [
1,
2,
3,
4]. In this context, low cardiorespiratory fitness (CRF, typically quantified as maximal oxygen uptake, VO
2max) has been documented to be a major risk marker of cardiovascular disease development and all-cause mortality, much stronger than other cardiometabolic risk factors, such as increased body mass index (BMI), smoking, hypertension, or elevated cholesterol levels [
5,
6,
7]. However, despite the overwhelming evidence for the beneficial health effects of being physically active, a large proportion of the adult population worldwide does not meet the recommended physical activity amounts (i.e., a minimum of 150 min of moderate or 75 min of vigorous-intensity aerobic physical activity per week [
8]) [
9] and this trend has been further amplified by the outbreak of the COVID-19 pandemic [
10,
11]. The individual reasons for physical inactivity are certainly diverse, but large-scale surveys conducted during the last decade on different continents have frequently revealed that “lack of time” constitutes a major barrier for regular physical activity and exercise [
12,
13,
14]. Thus, in recent years, the development and evaluation of more time-efficient exercise modalities has increasingly become a focus in the research field of exercise physiology and sports science [
15]. In this light, low-volume high-intensity interval training (LOW-HIIT) has emerged as a promising exercise type that has shown great potential in improving a variety of health outcomes with less time effort compared to more traditional exercise methods, such as (moderate-intensity) longer-distance endurance training [
16,
17,
18]. LOW-HIIT is typically characterized by short, vigorous exercise bouts separated by low-intensity recovery periods and involves, per the previous definition, a maximal total session duration of ≤30 min (including warm-up, recovery periods, and cool-down) [
17]. Previous research from our group has shown, for example, that LOW-HIIT can significantly improve VO
2max and various cardiometabolic markers in sedentary but otherwise healthy individuals [
19] and in obese metabolic syndrome patients [
20,
21,
22,
23] after only a few weeks.
However, in contrast to longer-distance endurance training or resistance exercise [
24], the potential of certain nutritional strategies and/or specific supplements to enhance the physiological adaptations in response to LOW-HIIT is still relatively sparsely investigated. It has been reported, for example, that there is evidence to suggest that several supplements, including sodium bicarbonate, nitrates, beta-alanine, caffeine and creatine, may provide potential benefits for improving HIIT adaptations [
16,
25]. Although it is generally accepted that athletes performing intense training, but also individuals engaged in recreational sports or general fitness routines, may require higher protein intakes compared to the sedentary population [
24], there is still a paucity of data regarding the impact of targeted post-session protein supplementation on HIIT-induced exercise adaptations [
25]. Previous research on the significance of proper protein intake is mainly based on studies using resistance training regimes, but it has also been documented that the consumption of a high-quality protein in doses of 20–40 g after a session of higher-volume endurance training may maximize muscle protein synthesis and support recovery processes [
24]. The physiological adaptations occurring after endurance exercise include the formation of new capillaries, mitochondrial proteins and other proteins involved in oxygen transport (e.g., hemoglobin and myoglobin) [
26,
27]. These adaptations play a critical role in the development of CRF (in addition to cardiac and pulmonary adaptations) and may translate into improved exercise performance over the longer term [
28,
29]. Thus, for these adaptation processes to run optimally, individuals participating in cardiovascular training modalities may hypothetically benefit from a post-exercise supplementation in the form of protein. However, given that most of the previous studies investigating the effects of protein supplements in conjunction with endurance training have typically used higher exercise volumes, there is currently still a lack of data on whether individuals engaged in LOW-HIIT may also benefit from a targeted post-session protein supplementation. Given the constantly high popularity of (LOW-)HIIT in recent years [
30], it appears timely to address this research gap.
Therefore, the present study aimed to investigate the impact of an eight-week LOW-HIIT program combined with a post-session intake of 40 g of whey protein in comparison to LOW-HIIT combined with an isocaloric placebo on CRF and cardiometabolic markers in a cohort of previously sedentary, healthy individuals. As HIIT has been shown to be a particularly effective training method to increase mitochondrial biogenesis and protein levels of glucose, lactate, and fatty-acid transporters in the skeletal muscle [
31], we hypothesized that targeted protein supplementation after LOW-HIIT would have beneficial effects on exercise-induced improvements in CRF and cardiometabolic outcomes.
4. Discussion
To the best of our knowledge, the present study was the first to investigate the influence of a targeted post-session protein supplementation on physiological adaptations to LOW-HIIT in a cohort of previously sedentary individuals. The main findings were as follows: (i) As expected, eight weeks of LOW-HIIT improved VO2max and some cardiometabolic markers, including BP and liver enzymes, but (ii) contrary to our hypothesis, the post-session supplementation of 40 g whey protein did not promote more favorable training adaptations compared to the placebo ingestion.
The average increase in VO
2max following LOW-HIIT (~3.2 mL/kg/min) was in accordance with the values observed in previous studies from our group, involving healthy sedentary individuals and obese metabolic syndrome patients (ranging from 3.0 to 7.1 mL/kg/min) [
19,
20,
21,
22,
23]. Given that VO
2max is a key indicator of health [
5,
6,
7], with each 1 mL/kg/min increase being linked to a reduction in CVD-related mortality by 9% [
46], this finding can be considered highly clinically meaningful. Notably, since a lack of time is one of the most frequently reported obstacles to participating in regular exercise, it is an important confirming result of this study that a LOW-HIIT protocol requiring less than 30 min per session can effectively improve CRF within only a few weeks. A decrease in BP following LOW-HIIT has been a consistent finding in the literature [
47], including research from our lab [
20,
21,
22,
23], and may be associated with exercise-induced improvements in vascular function [
48]. The average reduction in systolic BP by ~8 mmHg that was observed after the eight-week LOW-HIIT period is very likely to provide a clinically meaningful benefit, since each 5 mmHg decrease in systolic BP has been suggested to lower the risk of developing major cardiovascular events by ~10%, even at normal or high–normal baseline BP values [
49]. In this context, we note that the average pre-intervention BP of our participants was in the normal range (i.e., ~127/81 mmHg), pointing to a powerful preventive anti-hypertensive effect of LOW-HIIT. Additionally, the significant impact of LOW-HIIT on some liver enzymes detected in the present study is in line with our previous findings showing improvements in ALT serum concentrations and the non-alcohol fatty liver disease (NAFLD) fibrosis score in obese metabolic syndrome patients, who performed 12 weeks of LOW-HIIT [
23]. Furthermore, the significant reduction (PLA-HIIT group,
Table 5) or trend towards a reduction (PRO-HIIT,
p = 0.067) in serum LDL levels following LOW-HIIT is also in accordance with a previous meta-analysis reporting that HIIT-based exercise interventions can have beneficial effects on the lipid profile [
50].
When it comes to improving exercise performance and maximizing adaptations to training, respectively, a large variety of nutritional supplements are frequently used by athletes and recreationally active individuals, including, for example, carbohydrates, protein, vitamins, minerals, and creatine. While protein is certainly one of the most popular supplements consumed by individuals involved in resistance training in order to promote gains in skeletal muscle mass [
24,
51], proper carbohydrate intake is undoubtedly the crucial nutritional strategy for maintaining blood glucose levels during prolonged endurance exercise and for replacing depleted muscle glycogen stores after exercise [
24,
52,
53]. Although the role of protein supplementation in endurance-training adaptations has been comparatively little examined, it has been hypothesized that individuals performing endurance-type training programs may also benefit from additional protein, particularly when consumed after exercise [
27]. These suggestions are mainly derived from a small number of studies, showing that post-endurance-exercise protein ingestion may increase myofibrillar synthesis and/or activators of mitochondrial biogenesis such as peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) [
54,
55,
56]. Given that mitochondrial function and capacity are closely related to CRF [
57] and cardiometabolic health [
58,
59], the potential beneficial impact of post-exercise protein ingestion on maximizing the training-related adaptations of the mitochondria, as detected in previous studies, is encouraging. However, it has also been pointed out that the evidence of the role of protein supplementation on endurance-training adaptations is still rather inconclusive. The current knowledge is limited by the lack of longer-term interventions (i.e., >6 weeks) and the fact that previous studies mainly focused on measuring muscle protein synthesis and biochemical endpoints (e.g., specific enzymes regulating mitochondrial biogenesis), which certainly contributed to a better understanding of the underlying physiological mechanisms but did not necessarily translate into proof of improved CRF and performance outcomes [
27].
In contrast to our hypothesis, we did not detect any beneficial effect of a post-session protein supplementation on changes in CRF and cardiometabolic health measures following eight weeks of LOW-HIIT when compared to a placebo, which could be due to several factors. First, it is important to consider that previous studies that observed greater gains in VO
2max when training was combined with a post-session protein intake all used a substantially higher training volume (typically continuous, longer-duration aerobic exercise of 45–60 min duration/session) [
54,
60,
61], while our LOW-HIIT protocol required as little as 14–24 min/session. Consequently, the total exercise volume of LOW-HIIT may simply not have been enough to cause an increased need for additional protein, and thus limiting the impact of the post-session supplementation. Second, both groups in the present investigation consumed adequate amounts of protein in their daily diet, which could have masked the potentially beneficial effects of protein supplementation. Third, it has been reported that repeated ingestions of 20 g protein during the post-exercise recovery period appear to be more effective compared to a single administration, at least when it comes to stimulating muscle protein synthesis after a resistance training session [
62]. Accordingly, in two previous studies that observed greater VO
2max improvements when endurance training was combined with protein supplementation, the supplements were ingested repeatedly. More specifically, a first dose was administered immediately after the termination of exercise and another either 1 h later [
55] or before sleep [
60], respectively. Thus, it might be speculated that in the present study, a greater effect could have been achieved with a different protein-feeding pattern (i.e., repeated ingestions during the post-exercise period instead of a single administration). However, for compliance and monitoring reasons, we decided to administer only a single post-session protein dose, which may also be more representative of the supplementation habits among most individuals engaged in exercise routines in the real world. Moreover, it has previously been demonstrated that a single ingestion of 40 g whey protein following whole-body resistance exercise resulted in a significant (and, compared to 20 g, greater) stimulation of myofibrillar protein synthesis in healthy individuals [
44]. Fourth, the self-reported daily carbohydrate intake among study participants was in the lower range of the recommendations for individuals involved in an exercise program [
24], which could have mitigated improvements in the study outcomes. However, daily carbohydrate intake was similar between PRO-HIIT and PLA-HIIT and would therefore affect both groups in a similar manner. Fifth, previous research indicated that the improvement in VO
2max after a period of six weeks of endurance training was mainly attributed to an increase in cardiac output and blood oxygen-transport capacity, while skeletal muscle adaptations (i.e., capillarization, mitochondrial volume density) contributed less to VO
2max adaptations [
63]. Thus, it is conceivable that an exercise intervention over a longer period would have been necessary to reveal more benefits of protein supplementation to changes in physiological variables involved in CRF and cardiometabolic measures. Taken together, our findings suggest that a single ingestion of 40 g whey protein immediately ingested after each session does not seem to augment adaptations of CRF and cardiometabolic measures in response to eight weeks of LOW-HIIT in previously sedentary healthy individuals.
There are some limitations that should be considered when interpreting the results of this study. First, we note that, except for the three days each at study entry and during the last intervention week, we did not control participants’ normal nutritional intakes and daily physical activities beyond the LOW-HIIT sessions. Moreover, we did not strictly standardize participants’ diet throughout the study period, with the exception of providing them nutritional recommendations at study entry in the form of handouts and recipes. Thus, although there were neither significant differences in daily nutrition, daily physical activity or MET-hours between the two recording periods nor between both groups, we cannot rule out that potential within- or between-group variations in participants’ nutrition, habitual physical activities or physical demands at their jobs during the non-recorded days of the eight-week intervention period could have affected the adaptations to LOW-HIIT. However, we highlight that it was the aim of our investigation to explore whether a targeted post-session protein supplementation could maximize the effects of LOW-HIIT without fundamentally influencing our participants’ habitual daily nutrition. Second, some of our conclusions are based on self-reported food- and physical-activity-recording data. In this regard, it must be kept in mind that individuals rather tend to underestimate their food intakes and to overestimate their physical activity levels during recording periods and/or that the recording per se may (unconsciously) affect the eating and physical activity behavior [
64], although we suppose that the thorough explanation and briefing on how to record food intake and physical activity should have decreased the degree of potential errors. Third, we administered a fixed dose of 40 g protein after each LOW-HIIT session, and it could be argued that it would have been a better approach to match the supplement dosing to each participant’s body weight. We note, however, that it is not unusual that post-exercise protein dose recommendations are expressed as absolute values [
24] and other previous studies applying a supplementation strategy with fixed amounts observed significant effects on VO
2max [
60,
61] or myofibrillar muscle protein synthesis [
44,
54], for example. Fourth, we acknowledge that we focused on evaluating the effects of post-session protein ingestion on CRF and cardiometabolic measures, but we did not investigate any specific markers associated with mitochondrial adaptation or muscle capillarization. Thus, we cannot rule out that we overlooked some relevant differences in skeletal muscle adaptations between PRO-HIIT and PLA-HIIT. Finally, we note that three participating women (PRO-HIIT, N = 1; PLA-HIIT, N = 2) were using contraceptive pills, which could potentially have an influence on blood pressure [
65]. However, due to the very small number of women using contraceptive pills and the comparable distribution between both groups, we did not expect a meaningful impact on our results. Moreover, we highlight that the pre- and post-intervention examinations were standardized according to the individual menstrual-cycle phase of all female participants.
In spite of these limitations, this is, to our knowledge, the first double-blind randomized placebo-controlled study to investigate the effects of a targeted post-LOW-HIIT-session protein supplementation on longer-term adaptations of CRF and cardiometabolic outcomes in previously sedentary individuals.