Effects of Exercise Training Intensity and Duration on Skeletal Muscle Capillarization in Healthy Subjects: A Meta-analysis : Medicine & Science in Sports & Exercise

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Effects of Exercise Training Intensity and Duration on Skeletal Muscle Capillarization in Healthy Subjects: A Meta-analysis

LIU, YUJIA1,2; CHRISTENSEN, PETER M.3; HELLSTEN, YLVA2; GLIEMANN, LASSE2

Author Information
Medicine & Science in Sports & Exercise: October 2022 - Volume 54 - Issue 10 - p 1714-1728
doi: 10.1249/MSS.0000000000002955

Abstract

Skeletal muscle capillarization is a central determinant of oxygen and nutrient delivery, and removal of metabolites in skeletal muscle (1) with important implications for health (2–4) and exercise performance (5,6). Accordingly, muscle capillarization is essential for insulin sensitivity (2–4) and seems to be linked with both whole-body maximal oxygen uptake (7) and critical power (8); the physiological limit above which a progressive loss of muscle metabolic homeostasis occurs. Thus, increases in muscle capillarization are considered advantageous for both health and exercise performance. The level of physical activity influences muscle capillarization; sedentary behavior, immobilization and bed rest are associated with regressive changes in the capillary network in some (9–12) but not all studies (13–15), whereas pioneering work from the early 1970s revealed that physical activity promotes growth of new capillaries (16,17). A number of studies have since investigated the impact of different physical training interventions on capillary growth, and in sedentary subjects, a few weeks of training typically result in a 10%–30% increase in capillary to fiber ratio (C:F) (11,12,17). In addition, cross-sectional studies report that muscle capillarization can reach mean C:F values of ~3.0 in elite endurance athletes (18–20) being around twice as high compared with that of untrained subjects with typical mean values of ~1.5–2.0 (11,12,17). One training study in humans, with a high time resolution by frequent muscle biopsies, showed a progressive increase in C:F during an 8-wk training period (17). Although the study only included five subjects, there was a clear numerical increase in C:F after week 1 and significant changes after just 5 wk in untrained subjects. However, there is a need for knowledge on how aspects such as training intensity, training duration (in weeks), number of training sessions, and total training volume (total volume = minutes per session × sessions per week × weeks duration) influence skeletal muscle capillary growth. This knowledge is important when planning exercise training for populations ranging from sedentary individuals to world-class athletes, as well as for rehabilitation after immobilization or bed rest in patients and athletes.

The growth of new capillaries, angiogenesis, in skeletal muscle is a closely regulated process involving a large number of factors that either stimulate or inhibit the extension of the capillary network. These angiogenic factors are not included in the present review (for a thorough review, see Egginton (1)), but the physiological stimuli leading to upregulation and downregulation of these angiogenic factors are very important in the understanding of how various forms of physical activity can improve skeletal muscle capillarization. Extensive studies from the research groups of Hudlicka and Egginton have revealed that physiological angiogenesis in skeletal muscle is effectively promoted by mechanical stimuli induced by the frictional force of flowing blood on the vascular endothelium, so-called shear stress, and passive stretch of vessels (21–24), but muscle metabolic signaling also seems to play a role (1,25). Although all three factors are stimulated during muscle activity, the magnitude of each may vary according to the intensity and duration of a given training session (26,27). To this end, there is a paucity of literature studying what types of activity are most effective in improving muscle capillarization, but recent evidence suggests that exercise of very high intensity does not induce angiogenesis to the same extent as exercise of moderate intensity (28–30). Because skeletal muscle capillarization is pivotal for both health and performance, specific knowledge about to what extent different exercise modalities induce capillary growth is warranted. Moreover, because inactivity caused by a sedentary lifestyle, bed rest, or immobilization after injuries may significantly reduce skeletal muscle capillarization (10–12), it is of utmost importance to establish evidence for how to efficiently induce angiogenesis for patients, “normal” people, and athletes.

The aim of this systematic review and meta-analysis was to interpret the efficacy of various training characteristics, including training intensity, duration, number of training sessions, and total training time, on muscle capillarization in healthy young and age individuals.

METHODS

This review was conducted and reported by following the recommendations and the checklist of the PRISMA (31). The review was registered with The International Prospective Register of Systematic Reviews (PROSPERO 42020191790) on July 13, 2020.

Search strategy and selection criteria

We performed a systematic literature search in the databases PubMed and Web of Science. The terms used for the database searches were “exercise,” “training,” “physical activity,” “running,” “cycling,” “swimming,” “skiing,” “walking,” “jogging,” “rowing,” “soccer,” “stretch,” “climbing,” “capil*.” The search was limited to publication dates before June 17, 2021, and study in humans, and language was restricted to English. If full text was not available, a copy was retrieved via interlibrary loan or contact with the corresponding author. Relevant review articles were screened to identify any additional suitable studies for inclusion in the review.

Eligibility criteria

The present meta-analysis was limited to exercise training interventions using endurance (“aerobic”) exercise. The a priori inclusion criteria for study inclusion were as follows: 1) trials with healthy, normal weight (body mass index 18.5–25 kg·m−2) adults (>18 yr); 2) participants completed a period of no shorter than 2 wk of exercise training, and studies were only included if they included a clear description of the training protocol; 3) C:F and/or capillary density (CD) in skeletal muscle reported; 4) results reported as a mean with an SD or SEM, both before and after the intervention; and 6) research papers written in English.

Based on defined inclusion criteria, by analyzing titles, abstracts, and full texts of potentially relevant articles, two independent reviewers from the author list screened the articles to determine their eligibility. Disagreement was resolved by discussion, and if consensus could not be reached, the corresponding author was consulted. In articles where absolute changes in C:F/CD or SD/SEM were not reported, the corresponding author was contacted and original data were obtained when possible (32,33). In case of no reply from the authors, the article was excluded (34).

Data extraction

Two reviewers extracted data independently using a predefined data extraction form which all authors agreed upon. For each included study, data were extracted regarding the first author’s last name, year of publication, characteristics of participants, sample size, exercise intervention (category, intensity, frequency, duration, sessions, training time, and scheme), and mean values of preintervention and postintervention with corresponding SD or SEM. The primary outcomes considered in this systematic review were C:F and CD. The secondary outcome was maximal oxygen consumption (V̇O2max). To further interpret the relationship between capillarization and aerobic metabolism, we also extracted enzyme activity of citrate synthase.

Study quality and reporting

The quality and risk of bias of the included studies was evaluated in duplicate by two reviewers using a bias scale (35) and adjusted to the nature of exercise training studies, which all authors agreed upon. The quality of evidence for each specific result was based on 11 factors: 1) eligibility criteria specified, 2) randomization specified, 3) allocation concealment, 4) measurement of maximum rate of oxygen consumption, 5) blinding of assessor, 6) exercise training scheme and outcome measures assessed, 7) dropouts, 8) between-group statistical comparisons reported, 9) measures and variability for all reported outcomes measures, 10) activity monitoring in control groups, and 11) method of capillary measurement reported. Higher scores represent better study quality and reporting. Dropouts had a score of 2, between-group statistical comparisons reported a score of 3, and all other points had a score of 1 for each. Disagreements in scores between reviewers were resolved by a third party.

Data analysis

All analyses were carried out using STATA 12.0 to calculate the mean differences (MD). The MD of C:F and CD between baseline and postintervention were calculated and pooled using a random-effects model when heterogeneity was moderate or high, and a fixed-effects model was used when heterogeneity was low. A positive effect size indicated a beneficial effect for the experimental group.

Heterogeneity was determined by visual inspection of the forest plots and with consideration of the I2. For statistical significance for I2, values of 25% to ≤50% were considered low heterogeneity; 50% to ≤75%, moderate heterogeneity; and >75%, high heterogeneity. Testing for overall effect (z score) was regarded as significant at P < 0.05. Egger’s test and asymmetry of funnel plots were undertaken to assess publication bias. To test the robustness of the overall weighted effect sizes, a sensitivity analysis was conducted.

Subgroup analyses

Subgroup analyses were conducted to explore the impact of capillarization related to intensity (see discussion hereinafter), duration (2–4, 4–8, 8–12, and >12 wk), sex, age, number of training sessions (<20, >20), and total training time (<1000, 1000–2000, and >2000 min) in sedentary and trained subjects, separately.

Intensity subgroups were defined as follows using % V̇O2max, % maximal heart rate (HRmax), or % incremental-test peak power output (iPPO; often also termed Wmax, which is used interchangeably in the present study):

  • - Low intensity: <50% of V̇O2max, or <60% of HRmax, or walking
  • - Continuous moderate intensity: 51%–80% of V̇O2max, 60%–85% of HRmax, or 55%–80% iPPO
  • - High-intensity intervals: 81%–100% of V̇O2max or 80%–110% iPPO or >85% HRmax
  • - Sprint intervals: >V̇O2max or >110% iPPO

Studies of sedentary and trained subjects were divided into the same subgroups according to intensity (Table 1). Each study was carefully assessed to ensure that the right subgroup was used and specifically that the studies with trained subjects were not misplaced (e.g., 81% of V̇O2max is not considered “high intensity” for well-trained subjects). Using % V̇O2 or HRmax to define intensity is not without limitations form a physiological perspective. However, because this was used by the majority of all included studies, we choose to group studies this way, albeit the use of lactate threshold and critical power (or speed) would provide more details (67–69). Workloads below the lactate threshold are characterized by steady-state conditions and low blood lactate levels (<~2 mmol·L−1) and often termed moderate intensity. Workloads above critical power are characterized by an inability to sustain steady-state conditions with muscle and blood metabolite accumulation, fatigue development, and often reaching V̇O2max. Workloads below critical power and above lactate threshold are often termed “threshold” or “heavy-intensity” training. Lactate threshold and critical power typically occur around 50%–60% and 70%–80% of V̇O2max, respectively (67–69), in subjects with low or modest training background, which represents most of the subjects in the included studies (Table 1). However, in the four studies using athletes, lactate threshold and critical power (or speed) are expected to occur at a higher % V̇O2max than in less trained subjects (18,70). This could potentially lead to actual continuous threshold training being classified as interval training for these trained subjects. However, that was not the case because two of the studies with trained subjects classified in continuous moderate intensity did exercise between 70% and 85% Wmax or HRmax, whereas the remaining two studies performed sprint intervals.

TABLE 1 - Characteristics of participants, training, and outcomes.
Study n
(M/F)
Age (yr) Training Status Training Modality Intensity Intensity Subgroup Frequency (per week) Duration (wk) Duration Subgroup (wk) Training
Sessions
Training Time (min) Outcome Reported
C:F CD V̇O2max CS
Andersen and Henriksson (17) 5/0 21 Sed. Cycling erg. 80% V̇O2max CON 4 8 4–8 32 1280 + + +
Klausen et al. (11) 6/0 24 Sed. One leg erg. HR about 170 CON 3 8 4–8 24 1520 + + +
Svedenhag et al. (36) 8/0 23 Sed. Cycling erg. 60%–75% V̇O2max a CON 4 8 4–8 32 1280 + +
Svedenhag et al. (37) 8/0 23 ± 4 Sed. Cycling erg. 60%–75% V̇O2max CON 4 8 4–8 31.6 1264 + + + +
Wallberg-Henriksson et al. (38) 10/0 30 ± 6 Sed. Jogging Endurance CON 3 8 4–8 24 1080 + + +
Hoppeler et al. (39) 5/5 29 ± 5 Sed. Cycling erg. 90% HRmax INT 5 6 4–8 30 900 + + +
Rosler et al. (40) 10/0 31 ± 5 Sed. Cycling erg. 90%–95% HRmax INT 5 8 4–8 40 1200 + + +
Denis et al. (41) 8/0 22 ± 3 Active Cycling erg. 70%–80% V̇O2max CON 4 20 >12 80 4800 + +
7/0 62 ± 4 Active Cycling erg. 70%–80% V̇O2max CON 4 20 >12 80 4800 + +
Coggan et al. (42) 12/0 64 ± 3 Sed. Endurance training 60%–85% HRmax b CON 4 36–48 >12 144–192 6480–8640 + + + +
0/11 64 ± 3 Sed. Endurance training 60%–85% HRmax CON 4 36–48 >12 144–192 6480–8640 + + + +
Suter et al. (43) 12/0 39 ± 8 Sed. Jogging 75% V̇O2max CON 4 24 >12 96 2880 + + +
Hepple et al. (44) 9/0 68 ± 3 Sed. Cycling erg. 60%–70% HRreserve CON 3 18 >12 54 1620 + + +
10/0 68 ± 3 Sed. Cycling erg. 6%–70% HRreserve CON 3 9 8–12 27 810 + + +
Lampert et al. (45) 5/2 41 ± 13 Sed. Cycling erg. 90% of powermax CON 3 12 >12 36 1620 + + +
LaStayo et al. (46) 7/0 24 Sed. Cycling erg. Ecce. 54%–65% HRmax CON 2–4 8 4–8 28 810 + + +
6/0 24 Sed. Cycling erg. Conce. 54%–65% HRmax CON 2–4 8 4–8 28 810 + + +
Masuda et al. (47) 6/0 23 ± 2 Active One leg erg. 70% work ratemax CON 5 5 4–8 25 1500 + + +
Jensen et al. (48) 6/0 25 ± 3 Sed. One leg erg. 150% of leg V̇O2max S-INT 3–7 c 6.8 ± 3 4–8 29 435 + + +
6/0 25 ± 3 Sed. One leg erg. 150% of leg V̇O2max S-INT 3–7 4 2–4 14 210 + + +
7/0 24 ± 4 Sed. One leg erg. 90% of leg V̇O2max INT 3 4 2–4 12 180 + +
7/0 24 ± 4 Sed. One leg erg. 90% of leg V̇O2max INT 3 6 4–8 18 270 + +
Mourtzakis et al. (49) 6/0 23 ± 2 Active One leg erg. 70% work ratemax CON 5 5 4–8 25 1500 +
Ostergard et al. (50) 14/5 31 ± 5 Sed. Cycling erg. 70% of V̇O2max CON 3 10 8–12 30 1350 + + + +
19/10 33 ± 5 Sed. Cycling erg. 70% of V̇O2max CON 3 10 8–12 30 1350 + + + +
Gavin et al. (32) 6/0 24 ± 2 Sed. Cycling erg. 65% V̇O2max CON 4 8 4–8 28 1680 + + +
8/0 64 ± 6 Sed. Cycling erg. 65% V̇O2max CON 4 8 4–8 28 1680 + + +
Daussin et al. (12) 7/4 45 ± 10 Sed. Cycling erg. 61% Pmax CON 3 8 4–8 24 720 + +
Iaia et al. (51) 9/0 34 ± 5 Runners Running 93% ± 0.5% Speedall out S-INT 3.4 4 2–4 13.6 231 + + + +
Høier et al. (52) 7/0 25 Sed. One leg erg. 80 cycles per minute LOW 4 2 2–4 8 720 + +
7/0 25 Sed. One leg erg. 80 cycles per minute LOW 4 4 2–4 16 1440 + + +
Bangsbo et al. (53) 0/18 19–47 Sed. Running 82% HRmax CON 2 16 >12 29.5 1770 + + + +
Murias et al. (54) 7/0 22 ± 1 Sed. Cycling erg. 70% V̇O2max CON 3 12 4–8 36 1620 + + + +
7/0 69 ± 7 Sed. Cycling erg. 70% V̇O2max CON 3 12 8–12 36 1620 + + + +
7/0 22 ± 1 Sed. Cycling erg. 70% V̇O2max CON 3 6 8–12 18 810 + + + +
7/0 69 ± 7 Sed. Cycling erg. 70% V̇O2max CON 3 6 4–8 18 810 + + + +
Høier et al. (55) 14/0 32 Sed. Cycling erg. 60%–68% V̇O2max d CON 3 4 4–8 13 780 + + +
Høier et al. (30) 9/0 32 ± 6 Sed. Cycling erg. ∼64% V̇O2max CON 3 4 2–4 12 720 + + +
9/0 32 ± 6 Sed. Cycling erg. ∼117%–124% V̇O2max CON 3 4 2–4 13 312 + + +
Gouzi et al. (56) 11/12 62 ± 6 Sed. Cycling erg. HRventilatory threshold e S-INT 3–4 6 2–4 20 1800 + +
Cocks et al. (57) 8/0 21 ± 3 Sed. Cycling erg. 65% V̇O2max CON 5 6 4–8 30 1500 + + +
8/0 22 ± 3 Sed. Cycling erg. All-out S-INT 5 6 4–8 30 45 + + +
Baum et al. (58) 10/0 31 ± 5 Sed. Cycling erg. 9%–95% HRmax INT 5 8 4–8 40 1200 + + +
6/0 36 ± 6 Sed. Home-based Tr. 75% V̇O2max CON 4 24 >12 96 2880 + + +
Boushel et al. (33) 7/2 34 ± 6 Sed. Skiing ~60% HRmax CON 6 4–8 42 2520 + + +
Gliemann et al. (28) 3/2 34 ± 11 Runners Running 75%–85% HRmax CON 3 8 4–8 24 + + +
Gliemann et al. (59) 6/5 46 ± 3 Sed. Cycling erg. 80% HRmax CON 3–4 f 8 4–8 29 1740 + + +
van Ginkel et al. (60) 14/13 67 ± 2 Sed. Skiing CON 7 4–8 28 ~5985 + + +
Prior et al. (61) 7/5 65 ± 23 Sed. Running 50%–75% HRreserve CON 3 24 >12 72 3240 + + +
Raleigh et al. (62) 23/0 20 ± 2 Active Cycling erg. 170% W max g S-INT 4 4 2–4 16 176 + +
Hesketh et al. (63) 10/0 20 ± 3 Sed. Cycling erg. ~65% V̇O2max CON 3 6 4–8 18 2700 + +
Mitchell et al. (64) 10/0 23 ± 5 Athletes Cycling erg. Sprint S-INT 2 4 2–4 8 22 + + +
Skattebo et al. (65) 9 28 ± 5 Trained One leg erg. 7%–85% of W˙peak CON 3–4 6 4–8 21 1050 + + +
Islam et al. (66) 10/0 22 ± 2 Active Running 9%–95% HRmax S-INT 4 4 2–4 16 640 + +
10/0 22 ± 2 Active Running 7%–75% HRmax INT 4 4 2–4 16 640 + +
CON, continuous moderate intensity; Conce., concentric; Ecce., eccentric; Erg., ergometer; INT, high-intensity intervals; LOW, low intensity; M/F, male/female; Sed., sedentary; S-INT, sprint intervals; Tr., training.
a60% V̇O2max (first week), 70% V̇O2max (weeks 2–3), 75% V̇O2max (weeks 4–8).
b60%–70% HRmax (first 12 wk), 80%–85% HRmax (week 13 to end).
c3 (weeks 1–2), 4 (weeks 3–4), 5 (weeks 5–7).
d60% V̇O2max (weeks 1–2), 68% V̇O2max (weeks 3–4).
eHR at the ventilatory or dyspnea threshold.
f3 (weeks 1–3), 4 (weeks 4–8).
gPercent of watt maximum at V̇O2max.

Taken together, continuous training in the present study is thought to encompass both moderate and heavy (threshold) intensity enabling long duration of training sessions. High-intensity intervals are thought to encompass intensities between critical power and V̇O2max, and sprint interval is above V̇O2max, both causing rapid fatigue development and thus shorter training duration possible. Studies in which a mixture of training intensity was applied throughout a training week were not included in the analysis. It should also be noted that, from a clinical perspective, 60%–85% HRmax is considered “vigorous” and thus is included in the “continuous” training subgroup in the present study (71). Training status was retrieved using the included study definition for sedentary versus trained, and the group was categorized as untrained if a trial failed to report training status. In all cases, clear definitions of either training history and/or an objective performance parameter were carefully assessed. No studies fall in between sedentary and trained.

RESULTS

Study Selection

The search yielded 1506 records, with 76 studies eligible for full-text review. Of these, 57 trials in 38 studies met our eligibility criteria. Forty-seven and 50 trials measured C:F and CD before and after exercise training separately. Data on capillarization originated from muscle biopsies obtained from vastus lateralis, apart from one study using soleus (38) and one study using gastrocnemius (42). V̇O2max was reported in 33 studies. Ten of the studies included measurements of citrate synthase activity (33,37,38,42,47,50,51,53,54,61). The flow diagram of the search process can be found in Figure 1.

F1
FIGURE 1:
Flow diagram of the search process.

Participant Characteristics

A summary of participants and outcome of all trials is listed in Table 1. C:F was reported from 391 participants, in both males and females. Thirty participants in three studies were active trained individuals or athletes, and 361 were sedentary health people. CD was reported from 428 participants of both sexes. Of these, 54 trained regularly or were identified as athletes in five studies, and 374 were sedentary health people. Ages ranged from 20.0 to 68.3 yr, and weights ranged from 63.9 to 96.4 kg.

Intervention Characteristics

The exercise training modality was primarily ergometer cycling, but included are also studies of jogging (38,43), running (28,51,61), skiing (33,60), home-based training (58), one-leg cycling (11), and one-leg knee extensions (47–49). Training intensity was determined as percentage of V̇O2max, HRmax, maximum work rate, heart rate reserve, maximum work rate, or all-out speed. Based on the criteria of intensity (see Methods), we divided the studies into low-intensity, continuous moderate-intensity, high-intensity intervals and sprint interval subgroups. The frequency of training varied from two to five times per week. Training duration was ranged from 2 to 48 wk. The total number of sessions ranged from 8 to 192. The total training time ranged from 22 min by a sprint training protocol to 8640 min by continuous moderate training for 48 wk in total.

Risk of Bias

Based on the bias scale described previously, the results of quality and bias are presented in Supplemental Figure 1 (see Appendix, Supplemental Digital Content, Fig. S1, Assessment of the studies included, https://links.lww.com/MSS/C610). The predominant domains contributing to risk of bias were inadequate reporting of randomization, adverse events, and blinding of participants and of assessors. The Amylase-Periodic Acid Schiff staining method (72,73) was originally used to identify capillaries wherefore antibody was not reported in research papers published before 2006. Two studies identified capillaries by electron micrographs without use of antibodies (46,58). There was moderate risk of activity control bias, because of insufficient reporting of daily physical activity monitoring. Low risk represented in eligibility criteria, exercise attendance, and primary outcome comparison.

Meta-analyses

Forty-seven studies reported the data of C:F before and after a clearly defined training intervention. Overall meta-analysis showed a significant increase in C:F (MD, 0.33; 95% confidence interval (CI), 0.30–0.37; Fig. 2A) with exercise training, with moderate heterogeneity (I2 = 45.08%, P < 0.0001). In the 50 studies reporting CD, exercise training led to a significant increase in CD (MD, 49.76 capillaries per millimeter squared (cap·m−2); 95% CI, 36.91–62.61 cap·m−2; I2 = 68.82%; P < 0.0001; Fig. 2B).

F2
FIGURE 2:
Forest plot of overall meta-analysis. A, Overall effect of meta-analysis comparing C:F before with C:F after exercise training. B, Overall effect of meta-analysis comparing CD before with CD after exercise training. The square and black line for each study represents the MD and 95% CI. Size of the square represents the weight of each study. The dotted vertical line represents the mean treatment effect. The diamond denotes an overall treatment effect and 95% CI.

Subgroup Analyses

Intensity

C:F was significantly higher after training in sedentary participants, except for participants in the low-intensity subgroup (low intensity: MD, 0.28 capillaries per fiber (95% CI, −0.04 to 0.59 capillaries per fiber); continuous moderate intensity: MD, 0.34 capillaries per fiber; 95% CI, 0.27 to 0.40 capillaries per fiber; high-intensity intervals: MD, 0.43 (95% CI, 0.28 to 0.58); sprint intervals: MD, 0.49 (95% CI, 0.25 to 0.72); Table 2). The change amplitude of C:F was largest in the high-intensity interval subgroup (Figs. 3A, 3). In trained individuals, C:F was not different after training for any of the training intensities included (Table 2). See more detail in Figure 4.

TABLE 2 - Results of subgroup analysis.
Subgroup Analysis C:F (capillaries per fiber) CD (cap·m−2)
MD 95% CI I 2 (%) P % Weight MD 95% CI I 2 (%) P % Weight
Intensity
 Sedentary Overall 0.37 0.31 to 0.43 45.6 0.002 100 54.51 38.12 to 70.89 78.7 0.000 100
LOW 0.28 −0.04 to 0.59 0 0.525 2.85 17.79 −42.91 to 78.49 0 0.342 3.74
CON 0.34 0.27 to 0.40 13.9 0.248 63.32 45.78 28.97 to 62.60 72.0 0.000 72.96
INT 0.43 0.28 to 0.58 25.28 0.000 25.28 87.65 47.86 to 127.45 65.8 0.032 11.73
S-INT 0.49 0.25 to 0.72 8.56 0.138 8.56 77.91 23.56 to 132.26 70.0 0.010 11.58
 Trained Overall 0.08 −0.10 to 0.26 0 0.385 100 32.00 0.10 to 63.90 56.3 0.025 100
LOW N N N N N N N N N N
CON 0.07 −0.26 to 0.39 N 0.097 84.36 41.24 −5.42 to 87.90 64.1 0.025 60.69
INT N N N N N 29.00 −12.72 to 70.72 N N 16.51
S-INT 0.14 −0.32 to 0.59 0 0.642 15.64 7.15 −59.22 to 73.51 45.8 0.174 22.80
Duration
 Sedentary Overall 0.37 0.31 to 0.43 45.6 0.002 100 54.51 38.12 to 70.89 78.7 0.000 100
2–4 wk 0.48 0.30 to 0.67 27.0 0.223 10.98 51.03 21.73 to 80.34 21.9 0.255 16.73
4–8 wk 0.39 0.31 to 0.47 61.24 0.002 61.24 62.70 37.67 to 87.73 86.3 0.000 53.87
8–12 wk 0.28 0.12 to 0.45 0 0.717 9.95 28.84 2.75 to 54.92 0 0.508 13.99
>12 wk 0.28 0.16 to 0.41 32.9 0.189 17.84 48.99 28.91 to 69.08 0 0.490 15.41
 Trained Overall 0.08 −0.10 to 0.26 0 0.385 100 32.00 0.10 to 63.90 56.3 0.025 100
2–4 wk 0.14 −0.32 to 0.59 0 0.642 15.64 15.32 −22.03 to 52.66 26.8 0.255 39.31
4–8 wk 0.07 −0.10 to 0.26 N 0.097 84.36 3.81 −34.51 to 42.12 0 0.551 33.84
8–12 wk N N N N N N N N N N
>12 wk N N N N N 91.30 51.63 to 130.97 0 0.605 26.85
Sex
 Sedentary Overall 0.37 0.31 to 0.43 45.6 0.002 100 54.51 38.12 to 70.89 78.7 0.000 100
Male 0.39 0.33 to 0.46 33.1 0.037 73.91 61.39 40.66 to 82.12 82.9 0.000 73.93
Female 0.22 0.14 to 0.29 0 0.490 11.05 63.51 35.28 to 91.74 0 0.963 7.62
Both 0.37 0.22 to 0.52 42.0 0.227 15.04 23.74 3.01 to 44.47 0 0.945 18.46
 Trained Overall 0.08 −0.14 to 0.31 25.76 0.39 100 32.00 0.10 to 63.90 56.3 0.025 100
Male 0.14 −0.32 to 0.59 0 0.64 15.64 40.15 0.69 to 79.61 62.0 0.022 74.92
Female N N N N N N N N N N
Both 0.07 −0.26 to 0.39 N N 84.36 6.94 −38.09 to 51.97 9.0 0.295 25.08
Age (yr)
 Sedentary Overall 0.37 0.31 to 0.43 45.6 0.002 100 54.51 38.12 to 70.89 78.7 0.000 100
18–40 0.37 0.30 to 0.45 53.5 0.000 73.51 55.06 35.18 to 74.93 82.6 0.000 75.73
40–60 0.35 0.07 to 0.64 59.2 0.086 7.06 50.08 −26.21 to 126.37 0 0.496 2.88
>60 0.38 0.27 to 0.49 0 0.002 19.43 49.42 31.25 to 67.58 0 0.661 21.39
 Trained Overall 0.08 −0.10 to 0.26 0 0.385 100 32.00 0.10 to 63.90 56.3 0.025 100
18–40 0.08 −0.10 to 0.26 0 0.385 100 20.78 −6.70 to 48.26 31.5 0.188 86.15
40–60 N N N N N N N N N N
>60 N N N N N 101.00 46.92 to 155.08 N N 13.85
Training sessions
 Sedentary Overall 0.37 0.31 to 0.43 45.6 0.002 100 54.51 38.12 to 70.89 78.7 0.000 100
<20 0.45 0.36 to 0.54 36.5 0.084 21.32 41.06 19.94 to 62.18 20.6 0.241 26.76
>20 0.32 0.27 to 0.36 37.7 0.022 78.68 60.19 40.02 to 80.36 82.7 0.000 73.24
 Trained Overall −0.04 −0.27 to 0.20 0 0.617 100 32.00 0.10 to 63.90 56.3 0.025 100
<20 0.14 −0.32 to 0.18 0 0.642 15.64 15.32 −22.03 to 52.66 26.8 0.255 39.31
>20 0.07 −0.26 to 0.39 N N 84.36 41.24 −5.42 to 87.90 64.1 0.025 60.69
Training time (min)
 Sedentary Overall 0.37 0.31 to 0.43 45.6 0.002 100 54.51 38.12 to 70.89 78.7 0.000 100
<1000 0.43 0.32 to 0.54 64.7 0.000 42.97 63.13 32.15 to 94.11 86.6 0.000 39.52
1000–2000 0.36 0.29 to 0.44 0 0.780 38.89 51.13 32.49 to 69.78 41.7 0.027 44.78
>2000 0.27 0.13 to 0.41 48.2 0.086 18.14 42.82 22.97 to 62.67 0 0.857 15.70
 Trained Overall 0.08 −0.10 to 0.26 0 0.385 100 32.00 0.10 to 63.90 56.3 0.025 100
<1000 0.14 −0.32 to 0.59 0 0.642 15.64 15.32 −22.03 to 52.66 26.8 0.255 39.31
1000–2000 0.23 −0.14 to 0.50 N N 42.58 17.32 −32.91 to 67.56 0 0.468 21.01
>2000 N N N N N 91.30 51.63 to 130.97 0 0.605 26.85
NA −0.10 −0.38 to 0.18 N N 41.78 −15.00 −74.25 to 44.25 N N 12.83
N, none no studies was included in this sub-analysis; NA, not mentioned referring to the studies that did not report training time.

F3
FIGURE 3:
The absolute mean change and relative (%) change of C:F and CD expressed as function of training intensity (A–D) and duration (E and F). Each dot represents a mean outcome of one study including sedentary (blue), trained (red), or age (green) subjects. White bar represents participants age 18–40 yr, and gray bar represents participants age >60 yr (E–H). A, Change of C:F in different intensity subgroups. B, Percentage of change of C:F in different intensity subgroups. C, Change of CD in different intensity subgroups. D, Percentage of change of CD in different intensity subgroups. E, Change of C:F in different duration subgroups. F, Percentage of change of C:F in different duration subgroups, G, Change of CD in different duration subgroups, H, Percentage of change of CD in different duration subgroups.
F4
FIGURE 4:
Subgroup analysis of C:F by intensity. The square and black line for each study represents the MD and 95% CI. Size of the square represents the weight of each study. The dotted vertical line represents the mean treatment effect. The diamond denotes an overall treatment effect and 95% CI. A, Forest plot of subgroup analysis of CD in sedentary subjects; B, Forest plot of subgroup analysis of CD in trained subjects.

CD was significantly higher in sedentary participants in the continuous-intensity subgroup and the high-intensity interval and sprint interval subgroups (continuous moderate intensity: MD, 45.78 cap·m−2 (95% CI, 28.97 to 62.60 cap·m−2); high-intensity intervals: MD, 87.65 cap·m−2 (95% CI, 47.86 to 127.45 cap·m−2); sprint intervals: MD, 77.91 cap·m−2 (95% CI, 23.56 to 132.26 cap·m−2)), but not in the low-intensity subgroup (MD, 17.79 cap·m−2 (95% CI, −42.91 to 78.49 cap·m−2)). The change in CD with training was largest after high-intensity interval protocols (Figs. 3C, D). CD was also higher after training in trained individuals, but the higher levels were not related to intensity (Table 2). See more detail in Supplemental Figure 2 (Appendix, Supplemental Digital Content, Fig. S2, Subgroup analysis of CD by intensity, https://links.lww.com/MSS/C610).

Duration of training intervention

Training increased C:F in sedentary participants for all training durations studied. The amplitude of change was larger when the training duration was short (< 8 wk) compared with long (>8 wk) (Table 2). Compared with young participants, C:F was increased more in age participants when training interventions was longer than 8 wk (Figs. 3E, F). In trained participants, training increased C:F, independently of training duration (Table 2). See more detail in Figure 5.

F5
FIGURE 5:
Subgroup analysis of C:F by duration. The square and black line for each study represents the MD and 95% CI. Size of the square represents the weight of each study. The dotted vertical line represents the mean treatment effect. The diamond denotes an overall treatment effect and 95% CI. A, Forest plot of subgroup analysis of C:F in sedentary subjects; B, Forest plot of subgroup analysis of C:F in trained subjects.

Training increased CD in sedentary participants for all training durations studied. The relationship between training duration and CD was inverse in young sedentary versus age participants; CD was increased most when duration ranged from 4 to 8 wk in young sedentary participants, whereas CD increased most after more than 8 wk in age participants (Figs. 3G, H). In trained participants, training increased CD only when training duration was longer than 12 wk (Table 2). See more detail in Supplemental Figure 3 (Appendix, Supplemental Digital Content, Fig. S3, Subgroup analysis of CD by duration, https://links.lww.com/MSS/C610).

Sex

Effect size of C:F in sedentary males was 156% of that in sedentary females (Table 2.). The effect size of CD was not different between sedentary males and females (Table 2). Currently, no studies have included trained female participants only. See more detail in Supplemental Figures 4 and 5 (Appendix, Supplemental Digital Content, Fig. S4, Subgroup analysis of C:F by sex (https://links.lww.com/MSS/C610) and Fig. S5, Subgroup analysis of CD by sex (https://links.lww.com/MSS/C610)).

Age

Exercise training increased C:F in each age groups of sedentary subjects with similar effect size (Table 2). Studies that measured C:F in trained participants included young participants only (Table 2). See more detail in Supplemental Figure 6 (Appendix, Supplemental Digital Content, Fig. S6, Subgroup analysis of C:F by age, https://links.lww.com/MSS/C610).

CD was not changed by training in sedentary subjects age 40–60 yr (Table 2). The effect size of CD in sedentary subjects age 18–40 yr was larger than those age >60 yr (Table 2). Training did not change CD in trained participants age 18–40 yr, but it was increased in trained subjects age >60 yr (only one study included in this subgroup). See more detail in Supplemental Figure 7 (Appendix, Supplemental Digital Content, Fig. S7, Subgroup analysis of CD by age, https://links.lww.com/MSS/C610).

Number of training sessions

In sedentary participants, the effect size of C:F (Table 2) was 40% larger when the training intervention involved less than 20 training sessions, compared with that more than 20 training sessions (Table 2). C:F was not changed after training in trained individuals in the range number of training sessions studied (Table 2). See more detail in Supplemental Figure 8 (Appendix, Supplemental Digital Content, Fig. S8, Subgroup analysis of C:F by number of training sessions, https://links.lww.com/MSS/C610).

The effect size of CD, when the training intervention has less than 20 training sessions, was 32% lower compared with interventions of more than 20 training sessions in sedentary participants (Table 2). CD was not changed after training in trained individuals in the range of training sessions studied (Table 2). See more detail in Supplemental Figure 9 (Appendix, Supplemental Digital Content, Fig. S9, Subgroup analysis of CD by number of training sessions, https://links.lww.com/MSS/C610).

Training time

In sedentary participants, exercise training improved C:F in the range of training time studied, and the effect size of C:F was larger after training interventions with <1000 min of training time compared with interventions with >1000 min of training time (Table 2). Exercise training did not change C:F in trained participants in the range of training time studied (Table 2). See more detail in Supplemental Figure 10 (Appendix, Supplemental Digital Content, Fig. S10, Subgroup analysis of C:F by training time, https://links.lww.com/MSS/C610).

In sedentary participants, CD was improved by exercise training, and the amplitude of change was largest when training time was longer than 1000 min. CD was increased only when training time was longer than 2000 min in trained individuals, but there were only two trials included (Table 2). See more detail in Supplemental Figure 11 (Appendix, Supplemental Digital Content, Fig. S11, Subgroup analysis of CD by training time, https://links.lww.com/MSS/C610).

Relationship between the Change of Capillarization and V̇O2max and CS

There was a positive correlation between the relative change of V̇O2max and C:F (R2 = 0.58, P = 0.002; Fig. 6A) but not CD (R2 = 0.004; Fig. 6B). The change in citrate synthase activity did not correlate to the change in C:F or CD (Figs. 6A, B).

F6
FIGURE 6:
A, Relationship between the training-induced change in C:F and relative maximal oxygen uptake (left y-axis, R-V̇O2max, P = 0.0002), and maximal citrate synthase activity (right y-axis, CS, P = 0.860). B, Relationship between the change in CD and V̇O2max (left y-axis, P = 0.077) and CS (right y-axis, P = 0.404).

Risk of Publish Bias

Funnel plots for C:F and CD of the analyses are presented in Supplemental Figures 12A and B (Appendix, Supplemental Digital Content, Fig. S12, Funnel plots for C:F from all of the included trials and for CD intervened by exercise from all of the included trials, https://links.lww.com/MSS/C610). The Egger’s regression test found no publication bias of C:F, whereas the funnel plots of CD were asymmetric, indicating potential publication bias. In the sensitivity analysis, with each study deleted from the model one by one, the results remained consistent across all deletions.

DISCUSSION

This meta-analysis provides the first comprehensive summary of training-induced changes in skeletal muscle capillarization based on studies from 1970 to 2022, including 57 trials from 38 studies with a total of 539 subjects. The main findings are that 1) to induce capillarization in untrained subjects, continuous moderate-intensity training (50%–80% of V̇O2max) and high-intensity interval training (>80% of V̇O2max) are more effective than training at low intensity (<50% of V̇O2max); 2) in untrained subjects, capillarization is induced after shorter training interventions of 2–4 wk with no further effect of longer interventions (8–48 wk) or more training sessions (>20 vs <20 sessions); and 3) in age subjects, longer training interventions (≥12 wk) are needed to induce capillarization. Finally, 4) capillary growth cannot be expected in already well-trained subjects within a time frame of training of 2–8 wk in duration.

Effect of exercise training on capillarization

The meta-analysis showed that the number of capillaries per muscle fiber and the density of capillaries are substantially increased by exercise training in previously untrained subjects. This was an expected outcome, as several studies have reported increased capillarization after a period of exercise training (30,48,52,55,58,62,66). Specifically, this meta-analysis provided data as to how much increase in capillarization can be expected after a training intervention of 8 wk or more: an increased capillary per muscle fiber ratio of ~24% or 0.33 cap/fiber (95% CI, 0.30–0.37 cap/fiber) and an increased CD of ~15% or 49.7 cap·m−2 (95% CI, 36.9–62.6 cap·m−2; Fig. 2). It is worth considering that C:F is generally the golden standard for determining growth of new capillaries, as the number of muscle fibers is not readily changed (74), whereas CD will be affected by a change in muscle fiber cross sectional diameter, which is often reported after a period of training. CD is, however, a better indicator of changes in diffusion distance for oxygen from blood to muscle fiber, and thus, changes in CD are the best measure of changes in blood to muscle diffusion capacity.

Training intensity

Subgroup analysis of studies using different training intensities showed a positive intensity-dependent response of changes in the capillary per fiber ratio. As such, the relative change in the capillary per fiber ratio was 21% higher after continuous moderate-intensity training compared with low-intensity training, and 54% higher after high-intensity interval training (Table 2). Interestingly, however, sprint interval training was not more effective than continuous or high-intensity interval training. This observation is maybe not surprising given that the accumulated workload and shear stress during sprints is considerably lower compared with the other training types (27). It may also be worth mentioning that sprint interval training has been shown to actually have a negative impact on angiogenic processes in already trained individuals (28,75). The reason for this remains to be determined, but it may be due to the fact that the short sprint intervals induce a less pronounced blood flow and thus shear stress response, compared with continuous moderate- and high-intensity interval training, and that total training volume is very low (27).

Training volume

Analysis of the time course of capillarization revealed that the change in capillarization occurs largely within the first few weeks of an exercise intervention. Moreover, the magnitude of capillary growth was not dependent on the total duration (in weeks) of the intervention, total number of training sessions, or total time spent on training. This finding concurs with findings in animals (76) and humans (48,55), where changes in muscle capillarization have been reported to plateau after the first weeks of training (17). In trained subjects, however, this early more marked growth of capillaries does not seem to take place with a change in training regime. Still, it was a surprising result that the studies characterized by a lower volume of training (<8 wk or <1000 min) had more significant increases in capillarization than studies with a higher volume of training (>8 wk or >2000 min; Table 2). Part of the explanation may be that relatively few studies in the present analysis were with a high volume, and thus, variability among subjects may have caused this finding. Because few studies had a long duration, implications for long-term development (in years) with regard to training of athletes should be made with caution. Considering that endurance athletes have a C:F of ~3 (18–20) and the average increase in C:F with training for 2–12 wk in the present meta-analysis of mainly untrained subjects is 0.33 from a starting point in C:F of ~1.7, it seems reasonable to assume that a slow growth of capillaries is happening during extended periods of training in athletes.

Combining the meta-analysis of training intensity and duration, an interesting observation becomes evident; that training intensity is more important than total training volume, if limited to a maximum duration of 12 wk. This has significant implications when designing exercise training interventions for healthy individuals, patient groups, and those in rehabilitation, where this new knowledge can be used to optimize training interventions based in the individual need and capabilities. It is important to state that more studies are still required to determine the optimal balance between intensity, frequency, and duration in different populations. In future studies, a high time resolution as used in the pioneering work by Andersen and Henriksson (17) is considered relevant to gain better insight of the temporal changes in capillarization (i.e., not just a pre and post sample). Based on the present findings, training intensity should be the main focus, within the limits of what is safe for the individual group of subjects and patients, but it is also important to consider that low training volume or duration per se should not be the ultimate goal considering all the other positive side effects of an active lifestyle.

Aging population

An important finding in this meta-analysis was that in subjects older than 60 yr, CD was increased more after interventions of longer durations, indicating that more training time is needed in this group before capillarization is evident (Fig. 3). However, the data also show that C:F increases as early as after 4–8 wk in some age subjects, similar to that of young subjects, suggesting that the angiogenic process is not delayed in age subjects. Instead, the lack of change in CD during the first weeks of training may be a result of the initial increase in muscle fiber area that may be more pronounced in the age population compared with a younger population. There are currently no studies in age individuals on the impact of shorter exercise training interventions (2–4 wk) on capillarization or on the impact of intensified training.

Training status

There is limited literature on the effects of a given training intervention on capillarization in trained individuals, but from existing literature included herein, it is clear that changes in capillarization are less apparent than in previously untrained subjects. This is likely a natural consequence of the high level of capillarization that is already present in well-trained subjects where the capillary network already has adapted sufficiently to the requirements of regular exercise (18,19).

With regard to the time course of change in muscle capillarization when exercise training is continued beyond the classic 8- to 12-wk interventions, there is again limited evidence available. There seems to be a rapid increase in the first few months, but it has also been shown in a cross-sectional study that compared with that at the age of 21 yr, elite road cyclists have a 35% higher CD at an age of 25 yr with similar fiber area, implying that the difference is caused by more capillaries around each fiber in the group with 4-yr longer training experience (77). This indicates that the angiogenic process does not stagnate but proceeds at a slow rate with years of training (77–79). Further support for angiogenesis occurring in trained subjects, albeit at slow rates per year, is the high C:F in endurance-trained elite athletes that can be 2 times that of average sedentary subjects (18,19). It cannot be excluded that a genetically determined high degree of capillarization is part of the explanation, but considering that baseline values in the present analysis in long-term trained subjects are 2.74 (95% CI, 2.61–2.86) compared with around 1.70 (95% CI, 1.68–1.72) in untrained subjects, it seems logical that training is a major reason for high C:F in endurance athletes.

Sex

The impact of sex on capillarization is severely understudied, and we were only able to include three studies where the effect of exercise training on capillarization was assessed in females compared with 40 studies in men. Interestingly, however, we find that changes in C:F with a period of exercise training were, on average, 56% higher in males compared with females (Table 2). The amount of data available does not allow for identification of an underlying mechanism, but it is well known that female sex hormones are important for angiogenesis in women (80,81). Menstrual cycle and menopausal status are therefore important considerations when studying training-induced angiogenesis in this group. Aspects regarding low weight of female studies (Table 2) and variability among subjects as discussed in the Training Volume section may also impact the conclusion. Clearly, more studies are warranted in female populations.

Relationship between changes of capillarization and V̇O2max

There was a positive correlation between changes in V̇O2max and C:F from the studies that reported both values. This observation confirms that the efficacy of a training intervention as assessed by a change in V̇O2max is related to the efficacy of the training intervention on capillarization. Accepting that an increase in cardiac output governed by high blood volume and stroke volume is key to an increase in V̇O2max during exercise with a large muscle mass (82), an increase in CD at the muscular level is considered a beneficial adaptation to accommodate oxygen transfer from capillaries to mitochondria. The present meta-analysis confirms that a training-induced increase in capillarization is paralleled by an increase in V̇O2max. The reader is referred to other articles discussing how capillary supply may or may not affect V̇O2max (83,84), but it is important to remember that, in healthy individuals, changes in V̇O2max are primarily determined by central factors at the level of the heart and the blood (85). From other studies, we also know that individuals with high V̇O2max also have greater enzyme activity of muscle citrate synthase being a marker of mitochondrial density (86). However, in the present analysis, the relative increase in muscle capillarization was not linked with a similar increase in muscle citrate synthase activity.

Bias

We included studies that reported capillarization before and after a training intervention, but they were not blinded randomized controlled trials. This is a given, as it is impossible to blind the participating subjects. Although it is possible to bind the investigators, this was not reported in most of the included studies and is a potential bias.

Strengths and limitations

In this meta-analysis, we have included studies with a single-mode, clearly defined training intervention, and thus, we have excluded studies with combined training, for example, where cycle training is combined with strength training (87–89). This limits the possibility to translate present findings to rehab programs and training interventions where strength training is also an important aspect. However, capillary growth is reported after mixed training in several of these studies (87–89).

Another important aspect that was not possible to assess in this study was the regression and regrowth of capillaries after injury or severe inactivity like bed rest or after rehabilitation (90–94). The rate at which capillary rarefaction occurs with inactivity and disease conditions is unknown, as is the potential and time frame required to regenerate capillaries. Studies of the time course of capillary rarefaction and growth in humans are warranted.

CONCLUSIONS

Based on the systemic review and meta-analysis, we conclude that exercise training improves skeletal muscle capillarization in previously sedentary subjects and that training at higher intensities increases the growth of capillaries the most, independently of training duration or volume. In age individuals, training interventions may need to be of longer durations to improve CD. In women, limited data are available, but indications are that the angiogenic process is slower compared with their male counterparts. Unlike in sedentary subjects exposed to training, already trained subjects do not experience an increase in the present analysis. Hence, little is known regarding optimal training to increase capillarization further in athletes.

Further studies addressing the time course of training-induced changes in capillarization in different populations and addressing capillarization in female and already trained participants are warranted.

All authors declare that they have no conflict of interest. The study received no funding.

The authors declare that the results of the study are presented clearly, honestly, and without fabrication, falsification, or inappropriate data manipulation. The results of the present study do not constitute endorsement by the American College of Sports Medicine.

Systematic review registration: PROSPERO 42020191790.

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Keywords:

CAPILLARIZATION; EXERCISE TRAINING; CAPILLARY TO FIBER RATIO; CAPILLARY DENSITY; META-ANALYSIS

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