Acute effects of apnea bouts on hemoglobin concentration and hematocrit: a systematic review and meta-analysis
Acute effects of apnea bouts on hemoglobin concentration and hematocrit: a systematic review and meta-analysis
APNEA AND HEMATOLOGICAL EFFECTS
ABSTRACT
Objective: This study aimed to systematically analyze the existing literature and conduct a meta-analysis on the acute effects of apnea on the hematological response by assessing changes in hemoglobin (Hb) concentration and hematocrit (Hct) values.
Methods: Research in Pubmed, The Cochrane Library, and Web of Science was carried out for studies in which the main intervention was voluntary hypoventilation, and Hb and Hct values were measured. Risk of bias and quality assessments were performed.
Results: Nine studies with data from 160 participants were included, involving both subjects experienced in breath-hold sports and physically active subjects unrelated to breath-holding activities. The GRADE scale showed a “high” confidence for Hb concentration, with a mean absolute effect of 0.57 g/dL over control interventions. “Moderate” confidence appeared for Hct, where the mean absolute effect was 2.45% higher over control interventions. Hb concentration increased to a greater extent in the apnea group compared to the control group (MD = 0.57 g/dL [95% CI 0.28, 0.86], Z = 3.81, p = 0.0001) as occurred with Hct (MD = 2.45% [95% CI 0.98, 3.93], Z = 3.26, p = 0.001).
Conclusions: Apnea bouts lead to a significant increase in the concentration of Hb and Hct with a high and moderate quality of evidence, respectively. Further trials on apnea and its application to different settings are needed.
Keywords: apnea; exercise; hematocrit; hemoglobin; hypoventilation; hypoxia
Key points: Acute apnea bouts significantly increase hemoglobin and hematocrit concentration.
- INTRODUCTION
Voluntary apnea involves a cessation of ventilatory mechanics for a certain period of time. Under non-pathological conditions, breath-holding leads to a series of physiological adaptions to optimize basic organic functions in the face of induced hypoxia and hypercapnia. The chemical expression of this decrease in oxygen saturation - hypoxemia - and the increase in CO2 concentration - hypercapnia - at the chemoreceptors is the main driver of the succession of these adaptative mechanisms. Still, their expression may also be enhanced by other adaptative dive reflexes commonly found in many species (1). Together, they effect a synchronous action of both sympathetic and parasympathetic nervous systems, translating into effects at different levels.
On the one hand, in the cardiovascular system, the most relevant consequences are bradycardia and peripheral vasoconstriction, which lead to a prioritization of blood supply to organs less resistant to anoxia, such as the heart and the brain (2). On the other hand, it has been shown that mainly the kidneys and, to a lesser extent, the liver, in the face of generalized hypoxia and lack of blood supply, secrete erythropoietin, a glycoprotein hormone that increases the maturation and production of red blood cells in the long term (3–5).
Simultaneously, but with more acute consequences on hematological values reflected in changes in hematocrit (Hct) and hemoglobin blood concentration (Hb), the splenic response to hypoxia has been studied. It is characterized by a contraction of its smooth musculature, causing the release of an extra blood volume of approximately 160 mL rich in erythrocytes, which increases the capacity for storage and transport of O2, with an increase in Hb of between 2.8% and 9.6% of total Hb (6,7). This addition of red blood cells to the system also has a buffering effect on blood and muscle pH, which inevitably acidifies due to the prioritization of anaerobic metabolism in the face of insufficient O2 in the periphery caused by the state of hypoxia and the consequent accumulation of lactate in the blood (8,9).
In practice, as well as in research, the organism can be subjected to a state of hypoventilation, defined as the significant reduction of ventilations per minute (10), through different types of interventions that vary in the effects provoked and their magnitude. In terms of lung volume, high lung volume protocols have been described, in which the increase in intrathoracic pressure accentuates the dive reflex by direct activation on sympathetic nervous system baroreceptors and hinders venous blood circulation. This contributes to a decrease in cardiac output and low lung volume, which seems to cause a greater difference in O2 concentration between arteries and alveoli and a consequent greater arterial O2 desaturation (11). This latter apnea training mode also seems to lead to greater glycolytic activity, reflected in a higher lactate concentration and an earlier onset of hypoxia, since in this case, hypercapnia is not a limiting factor and allows the establishment of hypoxemic conditions in the absence of a hypoxic environment (12). Differences can also be found as to whether apnea is static or dynamic. The latter would entail, in addition to the processes derived from the apnea itself, a series of physiological changes which, regardless of the movement parameters used, will tend toward a predominantly anaerobic metabolic profile due to the increase in hypoxemic stress induced. A recent systematic review with meta-analysis (13) conclude that apnea training might increase peak blood lactate concentration tolerance, potentially related to improved performance in sports requiring high anaerobic capacity, on the other hand, according this review, further evidence is required to analyze potential benefits of apnea training on aerobic capacity.
This translates into a greater increase in circulating erythropoietin levels when compared to static apneas (14). Although not determinant, stimulation of the pulmonary stretch receptors and the ophthalmic branch of the trigeminal nerve by immersing the face in water potentiates these responses to breath-hold, but mainly the cardiovascular over the hematological one, suggesting different activation pathways (15). Although the human response to apnea has been extensively studied, the hematological consequences seem to show differences in their evocation and presentation among the other effects of apnea, so it is not possible to understand all these mechanisms as indivisible and to generalize the same activation processes for all. This article aimed to systematically review the scientific literature and perform a meta-analysis to assess the acute effects of apnea on hematological response by assessing changes in Hb concentration and Hct.
2. METHODS
This review has been conducted following the guidelines of the different items outlined in the PRISMA statement for systematic reviews and meta-analyses (16).
2.1 Search strategy
The literature search was carried out during November and December 2021 in PubMed, The Cochrane Library, and Web of Science meta-search engines. The title, abstract and keyword search fields were searched as follows: “TITLE”: ("apne*" OR "apnoe*" OR "hypoventilation" OR "hypoventilatory" OR "breath hold*") NOT ("sleep apnea" OR "sleep apnea" OR "obstructive sleep apnea" OR "obstructive sleep apnea" OR "osa" OR "infant apnea" OR "infant apnea" OR "hypoventilation syndrome") AND “TITLE/ABSTRACT”: ("hemoglobin" OR "haemoglobin" OR "Hb" OR "hematocrit" OR "haematocrit" OR “Hct”). The search, identification and screening of studies was performed independently by the two reviewers (O.LR and L. SF). Discrepancies were resolved by a third independent reviewer (F.DAF). For each search, titles and abstracts were screened to exclude irrelevant records. All articles meeting the eligibility criteria were downloaded and citations and reference lists were checked for other eligible articles for the study. In addition, a parallel search was carried out in Open Grey and Grey Literature Database.
2.2 Inclusion criteria
Inclusion criteria were: (1) clinical study in healthy subjects; (2) study of at least one of the variables studied in this review; (3) main intervention was voluntary hypoventilation; (4) prospective analysis of variables; (5) study of acute effects. Studies were excluded if: (1) animal study; (2) choice of subjects due to a pathological condition; (3) study of short-, medium- and long-term effects; (4) studies that did not meet the inclusion criteria.
2.3 Risk of bias in individual studies
The risk of bias of each article was assessed using the Cochrane risk of bias assessment tool (17). The overall quality of each article included in this review was rated as “high risk of bias,” “unclear risk of bias” or “low risk of bias.” The domains assessed were: (1) random sequence generation (selection bias); (2) allocation concealment (selection bias); (3) blinding of participants and personnel (performance bias); (4) blinding of outcome assessment (detection bias); (5) incomplete outcome data (attrition bias); (6) selective reporting (reporting bias); (7) other bias. Two reviewers (O.LR and L. SF) independently assessed the risk of bias and disagreements were resolved by a third reviewer (F.DAF). Inter-rater reliability was analyzed by calculating the Kappa index.
2.4 Quality assesment
The quality of evidence was determined using the GRADE (Grading of Recommendation Assessment, Development and Evaluation) scale (18). The five factors of the scale were assessed: risk of bias; inconsistency (heterogeneity); indirectness (evidence addresses the review question); imprecision (width of confidence intervals); and publication bias (funnel plots). These points lead the variables studied to an evidence score: “High” confidence that the actual effect is close to the effect estimate; “Moderate” moderate confidence in the effect stimulus with the actual effect likely to be close to the effect estimate, but likely to be substantially different; “Low” confidence in the effect estimate is limited and the actual effect may be substantially different from the effect estimate; “Very low” confidence in the effect estimate and the actual effect is likely to be substantially different from the effect estimate.
2.5 Statistical considerations
Meta-analytical procedures were applied to assess the possible effects of apnea bouts on the variables studied. Despite the utilization of random allocation procedures, potential discrepancies in baseline measures were mitigated for the meta-analysis through the extraction of mean and SD values of post-pre intervention changes within groups, in accordance with the following formulas, considering r = 0.5:
The absolute mean difference (MD) with 95% confidence intervals (CI) between apnea training (AT) and control group (CG) was calculated using a random-effects model. Significance for the overall effect was set at p < 0.05. Heterogeneity of the analyzed studies was assessed by an I-squared test (I2). The significance level of the I2 test was set at p < 0.05; the I2 represents the proportion of effects that are due to heterogeneity versus chance (19). Within controlled trial studies, a positive effect indicates a greater improvement in AT compared to CG in Hb or Hct, while a negative effect means the opposite. Funnel plots (Egger’s test was not possible) were used to assess publication bias (17). All statistical analyses were performed with RevMan 5.3.2 (Nordic Cochrane Centre).
3. RESULTS
3.1 Final study selection
The search process yielded a total of 325 candidate articles for inclusion in the review. After excluding duplicates and adding one article selected through reference lists, a total of 178 initial results were available. Ninety-five of these were excluded from the review after screening via title and abstract against the exclusion criteria, leaving a total of 81 studies. Seven of these were excluded because the full manuscript was not accessible and one because it was a protocol, leaving 74 clinical trials. Finally, 68 of these were excluded as they did not meet the determined inclusion criteria, leaving a total of six trials. However, one study included four comparisons that could be analyzed independently, leaving a total of 9 studies to be included in the review and meta-analysis (20). Figure 1 summarizes the selection of studies using a flow chart.
- Figure 1 about here-
3.2 Characteristics of the participants
The total sample size analyzed in the review is 160 subjects, which were divided into hypoventilation training (82 participants) and control group (78 participants). The mean age of the subjects was 23.96 ± 8.36 years old. Participants included a variety of athletes such as swimmers (21), free-divers (20) and cyclists (22), and non-athlete participants (20,23–25).
3.3 Characteristics of the studies selected
The mean duration of the studies was seven days, and all apnea interventions were performed in a single session. The procedures of Yildiz et al. and (24) were performed in a single day, as the control group was different from the intervention group. The rest of the studies were crossover clinical trials. Elia et al., (20), Bouten et al., (25), and Sperlich et al., (22) left washout times of one week between apnea training and control intervention. Robertson et al., (21) set washout times in the protocol between two and seven days, but since two other conditions not relevant to the present review were analyzed, the actual time between apnea training and control intervention could have been a total of at least six days and, at most, 21 days. Woorons et al., (23) protocolized washout times of 72 hours, but also analyzed two other conditions not relevant to the present review that could have been performed on the same day as the apnea training and the control intervention.
Regarding the characteristics of the apnea intervention, Elia et al., (20) combined static and maximal dynamic apneas in the experimental group, being the only trial where they were performed underwater. Bouten et al., (25), Yildiz et al., (24), Robertson et al., (21) and Sperlich et al., (22) used maximal static apneas. Woorons et al. (23) used a hypoventilation training protocol characterized by inhalations every four seconds while cycling at 65% of VO2max.
All interventions consisting of more than one apnea protocolized two-minute inter-apnea rests. In the study of Woorons et al., (23) active breaks of one minute were taken between sets. Detailed characteristics of the interventions in each trial are shown in Table 1.
- Table 1 about here -
3.4 Level of evidence and quality of the studies
Regarding the risk of bias analysis, a high concordance was achieved between the two investigators in charge of data extraction, resulting in a Kappa index of 0.905 prior to correction of controversial findings. The risk of bias found in several of the studies was mainly attributed to the lack of allocation concealment to the different interventions and blinding of participants. Among the other sources of bias, a high percentage of moderate risk of bias was found.
The methodological quality of the trials as assessed by the GRADE scale was shown to be “High” for the plasma Hb concentration studied in a total sample of 160 subjects, with a mean absolute effect of 0.57 higher. It can therefore be assumed that there is high confidence in the agreement between the effect obtained in the studies and the estimated effect. For the hematocrit percentage analyzed in a total of 102 subjects, the methodological quality was defined as “Moderate,” with the I2 statistic showing some inconsistency in the findings with low heterogeneity among the six included studies (I2=20%). In this case, the mean absolute effect was 2.45% higher. For changes in hematocrit caused by apnea, it must be considered that the effect found may not correspond to the estimated one, so further studies on this outcome may yield relevant information about the effects of apnea on it. Summary information is provided in Table 2.
- Table 2 about here -
3.5 Hemoglobin
A total of 9 studies analyzed the effects of apnea on Hb (20–25). The overall quality of evidence was determined to be “High” for the Hb study (Table 2).
Hb increased to a greater extent in the apnea group compared to the control group (MD = 0.57 g/dL [95% CI 0.28, 0.86], Z = 3.81, p < 0.0001). The I2 statistic showed non-significant heterogeneity between the included studies (I2 = 0%). These results are shown in Figure 2a.
3.6. Hematocrit
A total of 6 studies looked at the effects of apnea on Hct (20,22,24). The overall quality of evidence was determined to be “Moderate” for the study of Hct (Table 2).
Hct increased significantly in the apnea group compared to the control group (MD = 2.45 % [95% CI 0.98, 3.93], Z = 3.26, p = 0.001). The I2 statistic showed a non-significant heterogeneity between the included studies (I2 = 20%). These results are shown in Figure 2b.
- Figure 2ab about here -
4. DISCUSSION
In the present review and meta-analysis of the acute effects of apnea on Hb concentration and Hct, a significant increase in both variables has been identified, raising the possibility of acute and momentary improvement of the hematological profile through this mechanism, with potential transfer to sports performance and the treatment of patients who take advantage of some of its therapeutic effects.
Given the current evidence, spleen contraction must be understood as the main precursor of this acute increase in Hb and Hct, as other hypotheses of increased diuresis or hematological concentration secondary to increased plasma extravasation have been ruled out in previous research as the main cause of this physiological response (6,26,27). In addition, Schagatay et al., (6) observed in 2001 that the acute increase in Hb concentration after successive apneas did not occur in participants who had undergone splenectomy as it did in those subjects who retained their spleen. Afterwards, in 2005, Schagatay et al., (8) observed that three maximal apneas were sufficient to evoke spleen contraction and a consequent increase in Hb and Hct, and determined this again in 2007, noting that the hematological response elicited by apnea developed over three to four successive apneas, unlike the peripheral vasoconstriction and bradycardia which manifested in full after the first 30 seconds of a single apnea and with a more short-term reversible character (28). In fact, Hb and Hct continued to increase in rest periods between successive apneas, which may be attributed in part to a delay due to blood circulation, but bradycardia and vasoconstriction returned to basal levels just after each apnea (28). Richardson et al., (29) observed in 2009 that the main stimulus for this contraction is at the level of chemoreceptors due to decreased blood O2 saturation and later in 2012 they identified a possible coactivating role of hypercapnia (30). It has also been suggested that hypercapnia may increase the release of sympathoadrenergic catecholamines which also appear to play an important role in evoking splenic contraction during exercise (31,32). In addition, the potentiation of the hematological response does not appear to be dependent on trigeminal ophthalmic branch stimulation, as is the cardiovascular response, manifesting in greater magnitudes of bradycardia and vasoconstriction when performing apneas by immersing the face in water (28).
As already pointed out by Schagatay et al., (6) in 2001, acute changes after repeated apneas depend on three main factors: the subject’s previous apnea training, the time at which the blood sample is taken and the duration of the apnea.
Adaptations caused by prolonged exposure to hypoxic conditions have been extensively studied in high mountain populations such as in Tibet, the Andes or Ethiopia, as well as in population groups whose lifestyle depends on the sea, such as the Ama, Asian pearl gatherers, or the Bajau, a nomadic sea tribe in Southeast Asia whose primary livelihood comes from underwater fishing (33–35). Hurford et al., (36) concluded that the magnitude of the increase in Hb and Hct after a series of five apneas was smaller in non-apneic subjects than in Ama divers. Similarly, chronic adaptations to apnea have been found in free divers, as in the study by Elia et al., in which it was concluded that elite free divers had a higher capillary density and a larger O2 reservoir both in blood, expressed as a higher concentration of erythrocytes, and in muscle, identified as a higher concentration of myoglobin (14). In addition, it has been shown that these subjects more exposed to hypoxia due to apnea have higher basal levels of Hb and Hct and that, although they do not have a larger splenic volume, the contraction of the spleen in these subjects seems to be more effective, leading to a greater increase in Hb and Hct than in subjects not trained in apnea and other groups of athletes such as cross-country skiers (31,37–45). Even so, apnea training for eight weeks does not seem sufficient to increase acute spleen contraction, although it does seem to increase spleen volume (46).
Another determinant of the acute changes in hematological profile after repeated apneas is the timing of blood sampling. Several studies have shown that, following an apnea, Hb and Hct values begin to fall within 3 minutes of apnea and, after 10 minutes, return to baseline values (6,8,29). In the articles included in this review, measurements were taken immediately after completion of the protocol. Only Sperlich et al., (22) and Bouten et al., (25), measured at 10 and 5 minutes respectively. However, Bouten et al., (25) observed that 10 minutes after the intervention the Hb and Hct values were slightly elevated without reaching their baseline values, which differs from other results previously obtained. Although there are differences between all the results reported, the trend indicates that the most relevant acute adaptations in Hb and Hct occur within 10 minutes after apnea, a fact that may be relevant in the practical application of this type of training.
The third and final determinant of the magnitude of these adaptations to apnea was proposed to be the duration of apnea, but it has been shown that there is no correlation between the duration of apnea and the increase in Hb, indicating that perhaps the effectiveness of splenic contraction, total erythrocyte volume and/or the genetic component are more important in the magnitude of the response (38).
The acute increase in Hb and Hct theoretically optimizes O2 storage and transport, so that performing apneas prior to sport or endurance exercise could increase the aerobic performance of subjects in the presence of apneas. In addition, the increase in erythrocytes also improves the buffering system in both muscle and blood, attenuating blood acidosis and oxidative stress (39). These characteristics of apnea, among others, mean that it should be considered as another tool when prescribing therapeutic exercise in patients due to its apparent capacity to affect their metabolic profile or even their performance, recovery and tolerance. During physical exercise, the cardiorespiratory capacity to supply oxygen to the muscle determines the VO2max value, so Hb plays a relevant role as a limiting variable of this aerobic performance (40). It has already been studied and proven in numerous studies that with a systemic increase in Hb after blood transfusion, the value of VO2max increases (41,42). Although these increases in Hb have been obtained by blood transfusion, according to the results of the present review, apnea bouts seem to offer the opportunity to achieve an increase in Hb autonomously, which could indirectly increase VO2 max. However, among the literature, most of the trials seem to indicate no positive effect of prior apnea on performance even when the increase in Hb is significant (21,22,24).
Regarding hematocrit, historically, there has been a tendency to increase the levels of this variable artificially to improve aerobic performance, reflected in an increase in VO2 max. However, under normal conditions, the increase in hematocrit values is a limiting factor for cardiac output due, among other things, to the direct relationship between hematocrit and blood viscosity, which can lead to a reduction in VO2max (47). In fact, it has been found that subjects with low hematocrit (<40%) have a greater aerobic capacity than those with higher hematocrit, who also have, as mentioned above, a higher blood viscosity (48). We commonly accept a direct relationship between Hct and Hb, so these findings make us wonder which of the two variables has greater relevance in aerobic performance and how great is the relevance of both within this, as it seems that there may be other variables that are of greater importance in this aspect. However, it has been observed that aerobic performance has remained constant even in cases of chronic red blood cell excess, indicating possible adaptive mechanisms (49). Doubts remain as to the degree of aerobic limitation posed by an increase in hematocrit because of its limiting effect on cardiac output due to the associated blood viscosity, so more quality studies are needed to identify the relationships between changes in hematocrit and hemoglobin after apnea training, and performance.
With regard to the transfer of apnea training to practice and according to the results obtained in the meta-analysis, dynamic apnea seems to generate a greater magnitude of hematological responses as identified in the trial by Elia et al. (pt.4) (20). In the absence of a swimming pool, the dynamic protocol could be adapted to perform apneas while walking or during any other mode of dynamic activity.
Limitations
The present review has some limitations that should be stated. Most of the included studies have been conducted with physically active subjects. They are, therefore, not transferable to untrained subjects or elite athletes since only Elia et al. and Field (20) analyzed an experienced apneic sample. These may be chronically adapted with physiological systems, such as oxygen transport, delivery, and utilization, already well optimized. Another limitation of this review is the difficulty in drawing solid conclusions about apnea protocols. The variety of intervention protocols for apnea training is a review to consider, as Table 1 shows. However, there does seem to be a tendency to give breaks of around two minutes, apneas vary in volume, frequency and whether they are dynamic or static.
Implication of findings.
Current high to moderate confidence evidence suggests that a single session of apnea bouts in physically active and healthy participants induces hematological responses, increasing hemoglobin concentration and hematocrit. These hematological responses, in congruence with cardio-respiratory mechanisms, facilitate breath-holding duration. To further solidify our understanding, future research should investigate these responses in sedentary individuals. By doing so, researchers will have sufficient and meaningful evidence to confidently establish the hematological responses after apnea bouts in healthy population. These findings may have practical implications for patients with respiratory or cardiovascular pathologies. The potential increase in hemoglobin concentration and hematocrit could be beneficial for improving respiratory function, decreasing fatigue, enhancing overall functioning, and ultimately reducing disability in these patients.
5. CONCLUSION
The systematic review and meta-analysis of the existing evidence on the acute effect of apnea bouts on Hb and Hct has found a significant increase in both variables, with a high quality of evidence for Hb and moderate for Hct; however, variations in the response to apnea in different subjects and the different outcomes that affect apnea mean that future trials are needed to investigate further the effects of apnea and its application to different settings.
Conflict of interest. Authors declare that they have no conflicts of interest relevant to the content of this review.
Funding. No sources of funding were used in the preparation of this article.
Authorship contribution. O.LR and L. SF performed the systematic searches, data extraction, risk of bias and methodological quality assessment and original draft writing. F.DAF contributed to the design and planning of the review, assisted as third assessor and performed the statistical analysis. J.FM contributed to data extraction, critically reviewed, edited, and prepared the final manuscript, tables and figures from the original draft. All authors approved the final submitted manuscript version.
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Figures and Table legends
Figure 1. Flow chart of information through the different phases of a systematic review.
Figure 2ab. Forest plot of the acute effects of apnea on (2a) hemoglobin concentration and (2b) hematocrit according to the results extracted in the meta-analysis. CI confidence interval, SD standard deviation, IV weighted mean difference. Review authors judgments about each risk of bias item presented as percentages across all included studies.
Table 1. Summary of the characteristics of the studies included in the review. Apnea training (AT), Control Group (CG), Hemoglobin (Hb), Hematocrit (Hct), No Divers (ND), Breath-hold Divers (BHDs)
Table 2. Grading of Recommendations Assessment, Development and Evaluation (GRADE) summary of finding.