Patients with the diagnosis of sickle cell disease (homozygote HbS, compound heterozygote HbS and HbC or β-thalassemia) were recruited from the Hemoglobinopathy Outpatient Clinic of the Hematology and Transfusion Medicine Center at the School of Medical Sciences, University of Campinas (UNICAMP).

Exclusion criteria comprised disorders that could prevent proper assessment or IMT during the study period, such as asthma or neurological disorders. For result analysis, patients who missed the reevaluation more than once or who remained for more than one week without training were also excluded. All patients were instructed to maintain their medical treatments throughout the research period.

This was a randomized study on inspiratory muscle training for SCD patients of true and sham load group (TG and SG respectively) design. All the patients who agreed to participate signed a written informed consent form. The study was approved by the UNICAMP ethics committee (Protocol number: CAAE - 44635415.2.0000.5404) and registered to http://www.ensaiosclinicos.gov.br website (registration number: RBR-6g8n92). Patients were randomized by order of evaluation, being sequentially alternated between SG and TG. The TG performed the IMT with a true load, whereas the SG performed the IMT with a device with no true load.

Both groups used the same equipment model for IMT and were unaware of which group they were in. The study began with an assessment that included spirometry and volumetric capnography (VCap) and assessment of respiratory muscle strength by maximal inspiratory and expiratory pressure, PImax and PEmax, respectively. Two scales, the Medical Research Council (MRC) dyspnea scale and the modified fatigue impact scale (MFIS) were then applied and the [Lac] was subsequently assessed before and after the 6-minute walk test (6MWT). After this initial assessment, the patients were instructed on how to use the IMT device and took it to train at home, returning every 6 weeks for PImax and PEmax assessment and training load adjustment. Each newly included patient received a training device with a true or sham load, according to an alternating list kept by the investigator. After a period totaling 18 weeks, the patients underwent the final evaluation composed of the same tests as the first evaluation (Figure 1).

Figure 1.

Patients flow diagram. Vcap, Volumetric capnography; PImax, maximum inspiratory pressure; PEmax, maximum expiratory pressure; MRC, Medical Research Council dyspnea scale; MFIS, Modified Fatigue Impact Scale; [Lac], blood lactate concentration; 6MWT, 6-minute walk test; Borg, modified Borg scale.

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The entire study was carried out at Hospital de Clínicas at UNICAMP. All evaluations at baseline and at the end of 18 weeks were performed by the same physiotherapists, according to protocols, and under the same conditions. The evaluations were carried out by three physiotherapists, one being responsible for volumetric capnography (VCap), the other for spirometry and the third, along with the second, for the other evaluations.

The pulmonary Function Tests (PFT) included spirometry and volumetric capnography (VCap) and were always performed in the morning at the Pulmonary Function Laboratory of the Pulmonary Diseases Service at the Hospital de Clinicas da UNICAMP. Spirometry is capable of measuring the inhalation and exhalation of air volumes as a function of time.15 All the patients underwent spirometry before and 20 min after the inhalation of 400 μg of salbutamol, using a spirometer (EasyOne-PC®, NDD Medizintechnik AG, Zurich, Switzerland). The test was performed according to the American Society guidelines15 and the analyzed values were: forced vital capacity (FVC), forced expiratory volume in the first second (FEV1), FEV1/FVC ratio and mean forced expiratory flow between 25–75% of expired vital capacity (FEF25–75%).

To perform the VCap, a CO2SMOS Plus 8100 (Dixtal/Novametrix; Respironics, Murrisville, PA, USA) was used. The VCap assesses the concentration of expired CO2 as a function of expired tidal volume, analyzing the functional components of tidal volume (VC) and dead space; this method enables the determination of the relationship between ventilation and perfusion.16–18

The Analysis Plus software was used for online recording of respiratory cycles, continuing for five minutes. An off-line sequence of patient respiratory cycles was selected at the end of the data collection to accommodate a variation of less than 15% for the expiratory tidal volume and less than 5% for the end tidal partial pressure of CO2 concentration (PetCO2). In addition, sequences showing phases 2 and 3 slopes equal to zero were eliminated, along with discrepant values.18

The PEmax and PImax were determined using a digital manuvacuometer (MVD30-Globalmed). The patient used a nasal clip and remained seated during these maneuvers. The PImax and PEmax evaluation maneuvers were performed from residual volume (RV) and total lung capacity (TLC), respectively.

For data analysis, the highest value was recorded, each maneuver being performed at least five times, provided that the difference between the highest and the lowest value among three values presented a difference less than 10%.19,20

The MRC scale consists of only five items, of which the patient chose the item that corresponded to how much dyspnea limited his or her daily living activities (DLA). The patient then reported the degree of dyspnea subjectively, choosing a value between 1 and 5, being: 1 (only suffers from shortness of breath during intense exercise), 2 (suffers from shortness of breath when walking hastily or climbing a ramp), 3 (walking slower than people of the same age due to shortness of breath or having to stop to breathe even when walking slowly), 4 (stops to breathe after walking less than 100 m or after a few minutes) and 5 (feels so short of breath that does not leave the house anymore, or feels short of breath when getting dressed). The higher the score, the greater the limitation of DLA due to dyspnea. When the patient reported no dyspnea status, the classification was 0.6 In Brazil, the MRC scale was validated by Kovelis et al.21

The MFIS consists of 21 items distributed in three domains: physical (9 items), cognitive (10 items) and psychosocial (2 items), so that for each item there are 4 possible answers, graded from 0 to 4, in which the highest scores reflect greater impact of fatigue.

For the physical domain, the scores vary from 0 to 36, the cognitive ranges from 0 to 40 and the psychosocial ranges from 0 to 8. Therefore, the total score of the MFIS is given by the sum of the three domains and ranges from 0 to 84 points. Values below 38 are classified as the absence of fatigue. From 38, the higher the score, the greater the degree of patient fatigue.

The MFIS underwent cross-cultural adaptation and was validated for the Portuguese language (in the Brazilian standard), by Pavan et al., in 2007.22

The [Lac] was measured pre- and immediately post-6MWT, using a portable lactate meter (Accutrend plus, Roche, Switzerland), which has a procedure previously validated in the scientific literature.23 Measurements were performed in blood collected from fingertips after the superficial perforation with a disposable lancet. The cleaning of the skin was carried out before and after the perforation with 70% alcohol-soaked cotton.

All the participants performed the six-minute walk test (6MWT) according to the standardized protocol24 before and after the IMT period. A pulse oximeter (Nonin Medical, Inc.; MN, USA) was used to evaluate pulse rate and oxygen saturation. The modified Borg scale (Borg) was used to evaluate dyspnea and perceived exertion rate scores before and after the 6MWT, ranging from 0 to 10 (0 being "nothing" and 10 being "maximal").25 The patients were gently and periodically encouraged to walk on a level surface 30 m long during the 6-minute test.

A specific inspiratory training device (POWERbreathe® Wellness, IMT Technologies Ltd., Birmingham, UK) was used for the IMT, with load adjustment based on PImax. Instructions for proper use at home consisted of exclusive diaphragmatic breathing, not using accessory respiratory muscles during training. The training group used an adjusted load of 30% of the PImax during the first two weeks of training, aiming to facilitate the adaptation to the use of the device, with the load being adjusted by 40% of the PImax during the third week, 50% of the PImax during the fifth week and 60% of the PImax during the sixth week of the IMT up to the end of the study. Every 6 weeks of IMT, respiratory muscle strength (PImax and PEmax) was reassessed and, if the PImax was altered, the load was readjusted. The SG group performed the training with the same device, but without the flap valve; however, load adjustments were simulated so that the patients did not realize that they had used a sham load. Patients from both groups were instructed to perform the IMT daily for 10 min per day with a 1-min rest after 5 min, 7 days a week, until completing 18 weeks of training. The number of repetitions was between 13 and 15 breaths per minute during the 10 min of daily training.

All participants received a paper notepad for notes on the date and time of the IMT. If they did not do IMT on a given day, they could also write down the reason that prevented them from doing so. In addition, for greater reliability when performing the IMT at home, patients were also contacted by cell phone (voice or text message) to verify that they were performing the training correctly and daily or to clarify any problems they might have had with the use.

Exploratory data analysis was performed through summary measures (median, minimum and maximum). Comparison between groups was performed using the Mann–Whitney, Fisher's exact or Chi-square tests. The comparison between times and groups was performed with the ANOVA for repeated measures with the transformed responses in ranks. Differences with a p-value < 0.05 were considered significant. All statistical analyses were performed using the Statistical Analysis System (SAS) for Windows, version 9.4 (SAS Institute Inc., Cary, NC, USA).

Regarding spirometry, none of the variables showed a significant change, comparing the pre- and post-IMT periods in both groups. However, FEV1, FEV1/FVC and FEF 25 - 75% showed a significant increase (p < 0.05) in the pre- and post-bronchodilator comparison, both in the initial and final evaluations, but not in the comparison between the evaluations (Table 2).

The results of volumetric capnography showed no statistically significant change (p > 0.05) in the variables evaluated in the comparison between the two groups (SG and TG), except for the inspiratory time (Ti) variable, which was significantly higher in the SG compared to the TG (p < 0.05, group effect). However, the following variables: inspiratory tidal volume (Vi), peak expiratory flow (PEF), phase 3 slope of the capnography curve (P3Slp), phase 3 slope normalized according to the expired tidal volume (P3Slp/Ve), peak inspiratory flow (PIF) and Tobin Index (RSBI), had a significant change regarding the time effect in both groups (p < 0.05) (Table 3).

In both groups, the PImax and PEmax significantly increased after the IMT (p < 0.05, time effect). For the PEmax, there was a significant increase in the second evaluation, in relation to the first, and in the fourth evaluation, in relation to the third evaluation. Regarding the PImax, there was a significant increase in the second evaluation, compared to the first, and in the third, in relation to the second (Figure 2A and B).

Figure 2.

A: Maximal inspiratory pressure (PImax) in the first, second, third, and last evaluation, PImax1 < PImax2 < PImax3, PImax4. B: Maximal expiratory pressure (PEmax) in the first, second, third, and last evaluation, PEmax1 < PEmax2, PEmax3 < PEmax4. Figure C: Medical Research Council (MRC) scale of dyspnea in the first and last evaluation, MRC1 > MRC2. Figure D: Distance measurements of the 6-minute walking test (6MWT) in the first and last evaluation, 6MWT1 < 6MWT2. The statistical test used was ANOVA on ranks transformation, showing a time effect of the training for both groups (p < 0.05). Statistical analysis showed no interaction or group effect. Values are median.

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The MRC scale showed a significant reduction in the final evaluation of the values measured (p < 0.05, time effect), the pre-IMT and for both the SG and TG (Figure 2C), with the medians of both groups varying in an identical manner. No significant changes were detected in the MFIS.

In both groups, there was a significant increase in the distance covered after the IMT, compared to the pre-IMT assessment (p < 0.05) (Figure 2D and Table 4).

The [Lac] was significantly increased (p < 0.05), comparing pre- and post-6MWT, before and after IMT in both groups; however, there were no changes in the comparison between the initial and final evaluations (Table 4).

The values reported by the patients on the Borg, used before and after the 6MWT application, showed a significant increase between the pre- and post-6MWT evaluations in both groups (p < 0.05), but there were no changes in the comparison between the initial and final evaluations (Table 4).

All the data were also analyzed in patients grouped according to the sickle cell disease severity, i.e., HbSS plus HbSβovs HbSC plus HbSβ+, and revealed no significant difference.