The study received financial support from The Svend Andersen Foundation. The Centre for Physical Activity Research is supported by TrygFonden Grants ID 101390, ID 20045, and ID 125132. JPH is funded by HelseFonden and Copenhagen University Hospital, Rigshospitalet, while HLH is funded by the Beckett Foundation.

The Scientific Ethical Committee for the Capital Region of Denmark has previously approved the measurement of DL,CO/NO at rest, during exercise, and in the supine position in both healthy volunteers and patients with chronic obstructive pulmonary disease (COPD) at our institution (protocols H-20052659, H-21021723, and H-21060230).
NOTE: Before DL,CO/NO is measured during exercise, a dynamic spirometry, and a cardiopulmonary exercise test (CPET) must be performed. The dynamic spirometry is used for quality control of the individual DL,CO/NO maneuvers, while the CPET is used to determine the workload at which DL,CO/NO is to be measured during exercise. In patients with airflow limitation, notably due to obstructive lung disease, it may be advantageous to supplement the dynamic spirometry with a whole body-plethysmography to obtain a valid measure of vital capacity. A medical health check to rule out any known contraindications before initiating CPET is recommended17. Importantly, the CPET should be performed at least 48 h prior to the DL,CO/NO measurement obtained during exercise, as prior vigorous exercise may affect DL for up to at least 24 h18,19.
1. Dynamic spirometry
NOTE: Dynamic spirometry should be performed in accordance with the current clinical guidelines from the ERS and ATS20.
<ol>
	<li>Measure weight (to the nearest 100 g) and height (to the nearest 1 mm).</li>
	<li>Ask the participant to sit in an upright chair.</li>
	<li>Perform a dynamic spirometry during a forced expired maneuver to identify the forced expired volume in 1 s (FEV1) and forced vital capacity (FVC) of the participant, as described elsewhere20.</li>
</ol>
2. Cardiopulmonary exercise test (CPET)
NOTE: CPET should be performed in alignment with current clinical recommendations21.
<ol>
	<li>Adjust the cycle ergometer according to the participant&#39;s height and place a heart rate (HR) monitor on the chest.</li>
	<li>Place the participant on the cycle ergometer. Equip the participant with a mask connected to a metabolic measurement system, to measure ventilation and pulmonary gas exchange throughout the test.</li>
	<li>Instruct the participant to begin cycling at a self-selected pace &#8805;60 rounds per min (RPM) and perform a 5 min warm-up period at a submaximal workload based on self-reported activity level, daily fitness and disease status (e.g., 15-150 W).</li>
	<li>Increase the workload by 5-20 W every min until the participant reaches voluntary exhaustion. The increments should be based on the participant&#39;s current fitness level, so that the test is expected to terminate 8-12 min after the commencement of the incremental phase.</li>
	<li>Instruct the participant to avoid other vigorous exercise for the next 48 h.</li>
</ol>
3. Calibration of single breath diffusing capacity equipment
NOTE: It is necessary to calibrate flow sensors and gas analyzers to ensure that measurements are both valid and reliable. The exact procedure is manufacturer- and device-specific. The calibration&#160;procedure, including biological control, should be completed on each study day, and if less than one study day is executed per week, additional weekly calibrations should be performed. The experimental setup is shown in Figure 1.
<ol>
	<li>Open the software program on the computer, and an automatic warmup period of 50 min will be initiated to ensure sufficient temperature of the pneumotach.</li>
	<li>Make sure that the containers with the test gases are open (See Figure 1D).</li>
	<li>Perform a gas calibration by first connecting the sampling line from the pneumotach to the MS-PFT Analyzer Unit plug-in termed CAL (See Figure 1B).</li>
	<li>Initiate the gas calibration by selecting Calibration on the Home Page (See Figure 2A) and choose Gas calibration. Start the calibration by pressing Start or F1 (See Figure 2B).</li>
	<li>Attach the sampling line to the pneumotach when the gas calibration is fulfilled and accepted.</li>
	<li>Perform a volume calibration using a valid 3 L syringe. Initiate the volume calibration by selecting Calibration on the Home Page (See Figure 2A) and choose Volume calibration. Start the calibration by pressing F1, and follow the instruction provided by the software (See Figure 2C).</li>
	<li>Make sure that the inspiratory bag is connected to MS-PFT analyzer unit (See Figure 1C).</li>
	<li>Complete the calibration procedure by performing a biological control measurement at rest in the sitting position. This should be performed by a healthy non-smoker to ensure reliability of the method. If the given subject&#39;s week-to-week variation in DL,CO or DL,NO varies more than 1.6 and 6.5 mmol/min/kPa (5 and 20 mL/min/mmHg), respectively, the variation can be due to machine error and should be investigated further12, 22.</li>
</ol>
4. Preparation of the participant
<ol>
	<li>Calculate the desired workload from the prior CPET results for the chosen intensity (% of maximal workload (Wmax)) at which the DL,CO/NO will be measured.</li>
	<li>At least 48 h after the participant has performed the CPET, ask the participant to return to the laboratory to obtain the DL,CO/NO measurement during exercise.</li>
	<li>Measure the height (in cm to the nearest mm), weight (in kg to the nearest 100 g) and Hb from capillary blood (in mmol/L&#160;to the nearest 0.1 mmol/L) of the patient.</li>
	<li>On the Home Page of the program choose Patient &#62; New patient (See Figure 2A) and fill in the required data: Identification, Last Name, First Name, Date of Birth, Gender, Height, and Weight of the participant. Continue by selecting OK or F1 (See Figure 2D).</li>
</ol>
5. DL,CO/NO measurement during upright rest
NOTE: DL,CO/NO measurements are performed in accordance with current clinical recommendations from ERS task force12.
<ol>
	<li>On the Home Page choose Measurement &#62; NO Membrane diffusing (See Figure 2E).</li>
	<li>Start the automatic resetting of the software, to zero the gas analyzer for all tests gases and to initiate the mixing of the test gases in the connected inspiratory bag. Initiate the automatic resetting by pressing F1 (See Figure 2F).
	<ol>
		<li>The automatic resetting takes 140-210 s. Observe the instructions provided by the software&#160;to recognize when to initiate the measurement. It is important to initiate the measurement immediately when the software instructs to Connect patient.</li>
	</ol>
	</li>
	<li>Place the participant in an upright chair equipped with a nose-clip. Instruct the participant on how to perform the maneuver as described below.
	<ol>
		<li>Ask the participant to use the nose-clip and begin normal tidal respirations through a mouthpiece connected to the pneumotach. To ensure a closed system for the measurements, make sure that the participant&#39;s lips are kept closed around the mouthpiece.&#160;</li>
		<li>After three normal respirations, instruct the participant to perform a rapid maximal expiration to reach residual volume (RV).</li>
		<li>When RV is reached, immediately instruct the participant to perform a rapid maximal inspiration to total lung capacity (TLC), targeting an inspiratory time of &#60; 4 s. During the maximal inspiration, a valve opens, allowing the participant to inhale the gas mixture mixed with a known concentration of NO (800 ppm NO/N2) in an inspiratory bag just prior to the inhalation.</li>
		<li>Ask the participant to carry out a breath-hold of 5 (4-8) s at TLC. During the inspiration an inspired volume (VI) &#8805;90% of the FVC (or plethysmography-based vital capacity) with a 4-8 s breath-hold time is targeted23&#160;(Table 1).</li>
		<li>After the breath-hold, instruct the participant to perform a strong steady maximal expiration with no interruptions.</li>
		<li>After the maximal expiration ask the participant to let go of the mouthpiece and nose-clip. The software will then calculate DL,NO and DL,CO without any command.</li>
	</ol>
	</li>
	<li>Use verbal encouragement throughout the maneuver to ensure that the participant reaches RV and TLC. Assess the acceptability of the maneuver as per Table 1.</li>
	<li>Perform the maneuver again after at least a 4 min wash out period, and until two maneuvers fulfil the acceptability criteria (Table 1) or until a total of 12 maneuvers (see below) have been performed on the same session.</li>
	<li>The DL,NO and DL,CO are reported according to the criteria outlined in Table 2. We also recommend that breath-hold time, inspired volume, and alveolar volume as reported. Furthermore, the number of acceptable and repeatable maneuvers should be reported, and findings based on maneuvers that either do not fulfill the acceptability or repeatability criteria should be interpreted with caution.</li>
</ol>
6. DL,CO/NO measurement during exercise
NOTE: A timeline of DL,CO/NO measurements during exercise is provided in Figure 3.
<ol>
	<li>Place the cycle ergometer at a distance that enables the participant to breathe through the mouthpiece without having to change the cycling position. Increase the height of the equipment so that the measurements can be carried out with a correct working position on the bicycle (See Figure 2).</li>
	<li>Place the participant on the cycle ergometer and place a&#160;HR monitor on the chest. Instruct the participant to perform each maneuver as outlined in step 5.3.</li>
	<li>Instruct the participant to begin cycling for 5 min at a submaximal workload, as a warm-up prior to the measurement.</li>
	<li>Increase the workload to the target intensity while simultaneously starting the automatic resetting of the device by pressing F1 (see step 5.2). The automatic resetting takes 140-210 s, which is sufficient to ensure that the participant has reached steady state.</li>
	<li>When the automatic resetting is finished, turn the mouthpiece to the participant and perform a maneuver as described below while the participant continues cycling at the target intensity.
	<ol>
		<li>Follow the steps in steps 5.4 to 5.5. Assess acceptability and repeatability criteria (Table 1) at each workload, and report as for measurements during rest (see step 5.6 and Table 2).</li>
	</ol>
	</li>
	<li>After completion of the maneuver, remove the mouthpiece&#160;and decrease the workload to 15-40 W. Perform the active recovery phase for 2 min after which repeat steps 6.4 and 6.5. The 2 min of active recovery and the 140-210 s during the automatic resetting provides a sufficient washout period of 4-5 min.</li>
</ol>

Assessing Pulmonary Alveolar-Capillary Reserve During Exercise

The equipment and software presented in the article is not free of charge. None of the authors is associated with any company providing the license to the software. All authors declare no competing financial interests.

The protocol provides a standardized approach to the measurement of DL,CO/NO during exercise using the dual test gas single-breath technique. Since the obtained DL,CO/NO-metrics increase due to pulmonary capillary recruitment and distension, the method provides a physiologically meaningful measure of the alveolar-capillary reserve.
Critical steps in the protocol 
The method requires an exhalation to residual volume followed by an inspiration to total...

Exercise leads to a considerable increase in energy demand compared to the resting state. The heart and lungs respond by increasing cardiac output and ventilation resulting in an expansion of the alveolar-capillary bed, mainly the recruitment and distention of pulmonary capillaries1. This ensures a sufficient pulmonary gas exchange, which can be measured by an increase in pulmonary diffusing capacity (DL)2,3,4. The first attempts to measure DL during exercise date back more than a century5,6,7.&#160;The ability to increase&#160;DL from the resting state is often referred to as the alveolar-capillary reserve8, 9.
Experimentally, the relative contributions of alveolar-capillary membrane diffusing capacity (DM) and pulmonary capillary blood volume (VC) to alveolar-capillary reserve can be assessed by different methods, including the classical multiple fractions of inspired oxygen (<img alt="Equation 1" src="/files/ftp_upload/65871/65871eq01.jpg" style="margin: auto;" />) method10. An alternative technique that may be useful in this context is the dual-test gas method, in which the DL to carbon monoxide (CO) and nitric oxide (NO) (DL,CO/NO) are concurrently measured11. This technique was developed in the 1980s, and takes advantage of the fact that the reaction rate of NO with hemoglobin (Hb) is substantially greater than that of CO, such that the pulmonary diffusion of CO depends more on VC than does NO. Hence, the main site of resistance (~75%) to CO diffusion is located within the red blood cell, while the main resistance (~60%) to NO diffusion is at the alveolar-capillary membrane and pulmonary plasma12. The concurrent measurement of DL,CO and DL,NO thus permits the assessment of the relative contributions of DM and VC to DL12, where the change in DL,NO observed during exercise thus largely reflects the expansion of the alveolar-capillary membrane. An additional advantage of this method when obtaining measurements during exercise is that it involves a relatively short breath-hold time (~5 s) and fewer maneuvers compared to the classical <img alt="Equation 1" src="/files/ftp_upload/65871/65871eq01.jpg" style="margin: auto;" /> technique, where multiple repeated maneuvers with a standardized 10 s breath-hold are performed at different oxygen levels. Although <img alt="Equation 1" src="/files/ftp_upload/65871/65871eq01.jpg" style="margin: auto;" /> has recently been applied with a shorter breath-hold time and fewer maneuvers at each intensity13. Nevertheless, <img alt="Equation 1" src="/files/ftp_upload/65871/65871eq01.jpg" style="margin: auto;" /> only permits a total of six DL,CO maneuvers per session, whereas up to 12 repeated DL,CO/NO maneuvers can be performed without any measurable effect on the resultant estimates14. These are important considerations when obtaining measurements during exercise since both a long breath-hold and multiple maneuvers may be difficult to perform at very high intensities or in patient populations who experience dyspnea.
The present paper provides a detailed protocol, including theoretical considerations and practical recommendations on the measurement of DL,CO/NO during exercise and its use as an index of the alveolar-capillary reserve. This method is easily applicable in the experimental setting and permits the assessment of how diffusion limitation in the lungs may affect oxygen uptake in different populations.
Theory and measurement principles 
The DL,CO/NO method involves a single breath of a gas mixture with the assumption that the gases distribute equally in the ventilated alveolar space after inhalation. The gas mixture consists of several gases including an inert tracer gas. The dilution of the tracer gas in the ventilated alveolar space, as based on its fraction in end-expiratory&#160;air, can be used to calculate the alveolar volume (VA)15. The gas mixture also includes the test gas CO and NO, both of which are diluted in the ventilated alveolar space and diffuse across the alveolar-capillary membrane. Based on their alveolar fractions, their individual rates of disappearance (k), also termed the diffusing constant, from the alveolar space can be calculated. By convention, the DL for a test gas measured during a single-breath maneuver, is derived by the following equation16:
<img alt="Equation 2" src="/files/ftp_upload/65871/65871eq02v3.jpg" style="margin: auto;" />
where FA0&#160;is the alveolar fraction of the test gas (CO or NO) at the onset of the breath-hold of the individual DL maneuver, while FA is the alveolar fraction of the test gas at the end of the breath-hold, and tBH is the breath-hold time. DL is mechanically equivalent to the conductance of the test gas across the alveolar-capillary membrane, through plasma and the red blood cell interior to hemoglobin. It thus depends both on the conductance of DM&#160;and the so-called specific conductance of pulmonary capillary blood (&#952;), of which the latter depends both on the conductance of the test gas in blood and on its reaction rate with hemoglobin10. Given that the reciprocal of conductance is resistance, the total resistance to the transfer of a test gas depends on the following resistances in series10:
<img alt="Equation 3" src="/files/ftp_upload/65871/65871eq03v2.jpg" style="margin: auto;" />
These components may be distinguished by concurrently measuring the DL to CO and NO, because these have different &#952;-values, and their respective DL values&#160;thus depend differently on VC. The pulmonary diffusion of CO depends more heavily on VC than does NO, with the main site of resistance (~75%) to CO diffusion being located within the red blood cell12. In contrast, the main resistance (~60%) to NO diffusion is at the alveolar-capillary membrane and pulmonary plasma, because the reaction rate of NO with hemoglobin&#160;is substantially greater than that of CO. Hence, by concurrently measuring DL,CO and DL,NO, changes in both DM and VC will markedly affect the former, while the latter will depend much less on VC, thus permitting an integrative assessment of the factors that determine DL.
The reporting of DL,CO/NO metrics may be done using different units. Hence, the European respiratory society (ERS) uses mmol/min/kPa, whereas the American Thoracic society (ATS) uses mL/min/mmHg. The conversion factor between the units is 2.987 mmol/min/kPa = mL/min/mmHg.

<table><tbody><tr><td>HemoCue Hb 201+&amp;nbsp;</td><td>HemoCue, Br&amp;oslash;nsh&amp;oslash;j, Denmark</td><td>Unkown</td><td>For measurements of hemoglobin</td></tr><tr><td>Jaeger MasterScreen PFT pro (Lung Function Equipment)</td><td>CareFusion, H&amp;ouml;chberg, Germany</td><td>Unkown</td><td>For measurements of D&lt;sub&gt;LCO/NO&lt;/sub&gt;</td></tr><tr><td>Mouthpiece</td><td>SpiroBac, Henrotech, Aartselaar, Belgium</td><td>Unkown</td><td>Used together with the Lung Fuction Equipment. (dead space 56 ml, resistance to flow at 12 L s&amp;minus;1 0.9 cmH2O)&amp;nbsp;</td></tr><tr><td>Nose-clip</td><td>IntraMedic, Gentofte, Denmark</td><td>JAE-892895</td><td></td></tr><tr><td>Phenumotach</td><td>IntraMedic, Gentofte, Denmark</td><td>JAE-705048</td><td>Used together with the Lung Fuction Equipment</td></tr><tr><td>SentrySuite Software Solution</td><td>Vyaire&#039;s Medical GmbH, Leibnizstr. 7, D-97204 Hoechberg Germany</td><td>Unkown</td><td></td></tr><tr><td>Test gasses</td><td>IntraMedic, Gentofte, Denmark</td><td>Unkown</td><td>Concentrations: 0.28% CO, 20.9% O2, 69.52% N2 and 9.3% He</td></tr></tbody></table>

dual test gas pulmonary diffusing capacity measurement during exercise in humans using the single-breath method

The combined single-breath measurement of the diffusing capacity of carbon monoxide (DL,CO) and nitric oxide (DL,NO) is a useful technique to measure pulmonary alveolar-capillary reserve in both healthy and patient populations. The measurement provides an estimate of the participant&#39;s ability to recruit and distend pulmonary capillaries. The method has recently been reported to exhibit a high test-retest reliability in healthy volunteers during exercise of light to moderate intensity. Of note, this technique permits up to 12 repeated maneuvers and only requires a single breath with a relatively short breath-hold time of 5 s. Representative data are provided showing the gradual changes in DL,NO and DL,CO from rest to exercise at increasing intensities&#160;of up to 60% of maximal workload. The measurement of diffusing capacity and evaluation of alveolar-capillary reserve is a useful tool to evaluate the lung&#39;s ability to respond to exercise both in the healthy population as well as in patient populations such as those with chronic lung disease.

The protocol was implemented in 2021 and at the time of writing a total of 124 measurements during exercise (i.e., 51 in healthy volunteers and 73 in patients with COPD of various severities) had been performed. The maneuvers, as well as data on fulfilled acceptability and repeatability criteria, and the failure rate are all provided in Table 3.
Calculations 
As an example, calculations from a single DL,CO/NO maneuver are&#160;...

Watch this Scientific Journal Video about Author Spotlight: Integrating Alveolar-Capillary Reserve Measurements in Exercise Adaptation and Therapeutic Strategies at JoVE.com

Author Spotlight: Integrating Alveolar-Capillary Reserve Measurements in Exercise Adaptation and Therapeutic Strategies

This protocol presents a method to assess pulmonary alveolar-capillary reserve measured by combined single-breath measurement of the diffusing capacity to carbon monoxide (DL,CO) and nitric oxide (DL,NO) during exercise. Assumptions and recommendations for using the technique during exercise form the foundation of this article.

Dual Test Gas Pulmonary Diffusing Capacity Measurement During Exercise in Humans Using the Single-Breath Method

The combined single-breath measurement of the diffusing capacity of carbon monoxide (DL,CO) and nitric oxide (DL,NO) is a useful technique to measure ...

dual-test-gas-pulmonary-diffusing-capacity-measurement-during

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631 Views.  Copenhagen University Hospital - Rigshospitalet. Ons onderzoek richt zich op het mechanisme van verminderde longgasuitwisseling in verschillende ziektetoestanden en hoe kennis hiervan kan worden gebruikt voor diagnostische en doelgerichte behandelingen. We streven ernaar het begrip van acute inspanningsaanpassing te verfijnen door de alveolaire capillaire reserve te beoordelen door middel van diffusiecapaciteitsmetingen tijdens inspanning. Conventionele longfunctietesten, zoals dynamische spirometrie, en diffusiecapaciteitsmetingen tijdens ...

Video: Auteur in de schijnwerpers: integratie van alveolaire-capillaire reservemetingen in inspanningsaanpassing en therapeutische strategieën

In vivo Measurement of the Mouse Pulmonary Endothelial Surface Layer

The endothelial glycocalyx is a layer of proteoglycans and associated glycosaminoglycans lining the vascular lumen. In vivo, the glycocalyx is highly hydrated, forming a substantial endothelial surface layer (ESL) that contributes to the maintenance of endothelial function. As the endothelial glycocalyx is often aberrant in vitro and is lost during standard tissue fixation techniques, study of the ESL requires use of intravital microscopy. To best approximate the complex physiology of the alveolar microvasculature, pulmonary intravital imaging is ideally performed on a freely-moving lung. These preparations, however, typically suffer from extensive motion artifact. We demonstrate how closed-chest intravital microscopy of a freely-moving mouse lung can be used to measure glycocalyx integrity via ESL exclusion of fluorescently-labeled high molecular weight dextrans from the endothelial surface. This non-recovery surgical technique, which requires simultaneous brightfield and fluorescent imaging of the mouse lung, allows for longitudinal observation of the subpleural microvasculature without evidence of inducing confounding lung injury.

In vivo Measurement of the Mouse Pulmonary Endothelial Surface Layer

The endothelial glycocalyx/endothelial surface layer is ideally studied using intravital microscopy. Intravital microscopy is technically challenging in a moving organ such as the lung. We demonstrate how simultaneous brightfield and fluorescent microscopy may be used to estimate endothelial surface layer thickness in a freely-moving in vivo mouse lung.

The endothelial glycocalyx is a layer of proteoglycans and associated glycosaminoglycans lining the vascular lumen. In vivo, the glycocalyx is highly ...

Using Continuous Data Tracking Technology to Study Exercise Adherence in Pulmonary Rehabilitation

Pulmonary rehabilitation (PR) is an important component in the management of respiratory diseases. The effectiveness of PR is dependent upon adherence to exercise training recommendations. The study of exercise adherence is thus a key step towards the optimization of PR programs. To date, mostly indirect measures, such as rates of participation, completion, and attendance, have been used to determine adherence to PR. The purpose of the present protocol is to describe how continuous data tracking technology can be used to measure adherence to a prescribed aerobic training intensity on a second-by-second basis.
In our investigations, adherence has been defined as the percent time spent within a specified target heart rate range. As such, using a combination of hardware and software, heart rate is measured, tracked, and recorded during cycling second-by-second for each participant, for each exercise session. Using statistical software, the data is subsequently extracted and analyzed. The same protocol can be applied to determine adherence to other measures of exercise intensity, such as time spent at a specified wattage, level, or speed on the cycle ergometer. Furthermore, the hardware and software is also available to measure adherence to other modes of training, such as the treadmill, elliptical, stepper, and arm ergometer. The present protocol, therefore, has a vast applicability to directly measure adherence to aerobic exercise.

Pulmonary rehabilitation is widely recognized in the management of respiratory diseases. A key component to successful pulmonary rehabilitation is adherence to the recommended exercise training. The purpose of the present protocol is to describe how continuous data tracking technology can be used to precisely measure adherence to a prescribed aerobic training intensity.

Pulmonary rehabilitation (PR) is an important component in the management of respiratory diseases. The effectiveness of PR is dependent upon adherence to ...

A Method to Study the Impact of Chemically-induced Ovarian Failure on Exercise Capacity and Cardiac Adaptation in Mice

The risk of cardiovascular disease (CVD) increases in post-menopausal women, yet, the role of exercise, as a preventative measure for CVD risk in post-menopausal women has not been adequately studied. Accordingly, we investigated the impact of voluntary cage-wheel exercise and forced treadmill exercise on cardiac adaptation in menopausal mice. The most commonly used inducible model for mimicking menopause in women is the ovariectomized (OVX) rodent. However, the OVX model has a few dissimilarities from menopause in humans. In this study, we administered 4-vinylcyclohexene diepoxide (VCD) to female mice, which accelerates ovarian failure as an alternative menopause model to study the impact of exercise in menopausal mice. VCD selectively accelerates the loss of primary and primordial follicles resulting in an endocrine state that closely mimics the natural progression from pre- to peri- to post-menopause in humans. To determine the impact of exercise on exercise capacity and cardiac adaptation in VCD-treated female mice, two methods were used. First, we exposed a group of VCD-treated and untreated mice to a voluntary cage wheel. Second, we used forced treadmill exercise to determine exercise capacity in a separate group VCD-treated and untreated mice measured as a tolerance to exercise intensity and endurance.

Two exercise paradigms were tested on a newly developed chemically induced menopausal mouse model to examine the impact of menopause on exercise capacity and cardiac adaption to exercise.

The risk of cardiovascular disease (CVD) increases in post-menopausal women, yet, the role of exercise, as a preventative measure for CVD risk in ...

Investigation into Deep Breathing through Measurement of Ventilatory Parameters and Observation of Breathing Patterns

In this protocol, two deep breathing patterns were shown to 15 participants to determine an easy yet effective method of breathing exercise for future application in a clinical setting. The women in their twenties were seated comfortably in a chair with back support. They were fitted with an airtight mask connected to a&#160;gas analyzer. Three electrodes were placed on the chest connected to a wireless transmitter for relaying to the electrocardiograph. They executed a 5 min rest phase, followed by 5 min of deep breathing with a natural breathing pattern, terminating with a 5 min rest phase. This was followed by a 10 min intermission before commencing the second instruction phase of substituting the natural breathing pattern with the diaphragmatic breathing pattern. Simultaneously, the following took place: a) continuous collection, measurement and analysis of the expired gas to assess the ventilatory parameters on a breath-by-breath basis; b) measurement of the heart rate by an electrocardiograph; and c) videotaping of the participant&#8217;s thoracoabdominal movement from a lateral aspect. From the video capture, the investigators carried out visual observation of the fast-forward motion-images followed by classification of the breathing patterns, confirming that the participants had carried out the method of deep breathing as instructed. The amount of oxygen uptake revealed that, during deep breathing, the work of breathing decreased. The results from the expired minute ventilation, respiration rate and tidal volume confirmed increased ventilatory efficiency for deep breathing with the natural breathing pattern compared to that with the diaphragmatic breathing pattern. This protocol suggests a suitable method of instruction for assessing deep breathing exercises on the basis of oxygen consumption, ventilatory parameters, and chest wall excursion.

Here, we present a protocol to assess two deep breathing patterns of natural and diaphragmatic breathing for their effectiveness and ease of execution. Fifteen participants were selected, utilizing an electrocardiograph and expired gas analyzer for measurement of the ventilatory parameters, together with visual assessment by video capture of thoracoabdominal movement.

In this protocol, two deep breathing patterns were shown to 15 participants to determine an easy yet effective method of breathing exercise for future ...

Behavior

Determining The Electromyographic Fatigue Threshold Following a Single Visit Exercise Test

Theoretically, the electromyographic (EMG) fatigue threshold is the exercise intensity an individual can maintain indefinitely without the need to recruit more motor units which is associated with an increase in the EMG amplitude. Although different protocols have been used to estimate the EMG fatigue threshold they require multiple visits which are impractical for a clinical setting. Here, we present a protocol for estimating the EMG fatigue threshold for cycle ergometry which requires a single visit. This protocol is simple, convenient, and completed within 15-20 min, therefore, has the potential to be translated into a tool that clinicians can use in exercise prescription.

This protocol describes the electromyographic fatigue threshold which demarcates between nonfatiguing and fatiguing exercise workloads. This information could be used to develop a more individualized training program.

Theoretically, the electromyographic (EMG) fatigue threshold is the exercise intensity an individual can maintain indefinitely without the need to recruit ...

Assessment of Pulmonary Capillary Blood Volume, Membrane Diffusing Capacity, and Intrapulmonary Arteriovenous Anastomoses During Exercise

Exercise is a stress to the pulmonary vasculature. With incremental exercise, the pulmonary diffusing capacity (DLCO) must increase to meet the increased oxygen demand; otherwise, a diffusion limitation may occur. The increase in DLCO with exercise is due to increased capillary blood volume (Vc) and membrane diffusing capacity (Dm). Vc and Dm increase secondary to the recruitment and distension of pulmonary capillaries, increasing the surface area for gas exchange and decreasing pulmonary vascular resistance, thereby attenuating the increase in pulmonary arterial pressure. At the same time, the recruitment of intrapulmonary arteriovenous anastomoses (IPAVA) during exercise may contribute to gas exchange impairment and/or prevent large increases in pulmonary artery pressure.
We describe two techniques to evaluate pulmonary diffusion and circulation at rest and during exercise. The first technique uses multiple-fraction of inspired oxygen (FIO2) DLCO breath holds to determine Vc and Dm at rest and during exercise. Additionally, echocardiography with intravenous agitated saline contrast is used to assess IPAVAs recruitment.
Representative data showed that the DLCO, Vc, and Dm increased with exercise intensity. Echocardiographic data showed no IPAVA recruitment at rest, while contrast bubbles were seen in the left ventricle with exercise, suggesting exercise-induced IPAVA recruitment.
The evaluation of pulmonary capillary blood volume, membrane diffusing capacity, and IPAVA recruitment using echocardiographic methods is useful to characterize the ability of the lung vasculature to adapt to the stress of exercise in health as well as in diseased groups, such as those with pulmonary arterial hypertension and chronic obstructive pulmonary disease.

To assess the pulmonary diffusion and vasculature responses to exercise, we describe the multiple-inspired oxygen diffusion capacity technique to determine capillary blood volume and membrane diffusing capacity, as well as agitated saline contrast echocardiography to assess the recruitment of intrapulmonary arteriovenous anastomoses.

Exercise is a stress to the pulmonary vasculature. With incremental exercise, the pulmonary diffusing capacity (DLCO) must increase to meet the increased ...

Conducting Maximal and Submaximal Endurance Exercise Testing to Measure Physiological and Biological Responses to Acute Exercise in Humans

Regular physical activity has a positive effect on human health, but the mechanisms controlling these effects remain unclear. The physiologic and biologic responses to acute exercise are predominantly influenced by the duration and intensity of the exercise regimen. As exercise is increasingly thought of as a therapeutic treatment and/or diagnostic tool, it is important that standardizable methodologies be utilized to understand the variability and to increase the reproducibility of exercise outputs and measurements of responses to such regimens. To that end, we describe two different cycling exercise regimens that yield different physiologic outputs. In a maximal exercise test, exercise intensity is continually increased with a greater workload resulting in an increasing cardiopulmonary and metabolic response (heart rate, stroke volume, ventilation, oxygen consumption and carbon dioxide production). In contrast, during endurance exercise tests, the demand is increased from that at rest, but is raised to a fixed submaximal exercise intensity resulting in a cardiopulmonary and metabolic response that typically plateaus. Along with the protocols, we provide suggestions on measuring physiologic outputs that include, but are not limited to, heart rate, slow and forced vital capacity, gas exchange metrics, and blood pressure to enable the comparison of exercise outputs between studies. Biospecimens can then be sampled to assess cellular, protein, and/or gene expression responses. Overall, this approach can be easily adapted into both short- and long-term effects of two distinct exercise regimens.

To assess the influence of exercise intensity on physiologic and biologic responses, two different exercise testing protocols were utilized. Methods outlining exercise testing on a cycle ergometer as an incremental maximal oxygen consumption test and endurance, steady state submaximal endurance test are described.

Regular physical activity has a positive effect on human health, but the mechanisms controlling these effects remain unclear. The physiologic and biologic ...

Integration of Brain Tissue Saturation Monitoring in Cardiopulmonary Exercise Testing in Patients with Heart Failure

Cerebral hypo-oxygenation during rest or exercise negatively impacts the exercise capacity of patients with heart failure with reduced ejection fraction (HF). However, in clinical cardiopulmonary exercise testing (CPET), cerebral hemodynamics is not assessed. NIRS is used to measure cerebral tissue oxygen saturation (SctO2) in the frontal lobe. This method is reliable and valid and has been utilized in several studies. SctO2 is lower during both rest and peak exercise in patients with HF than in healthy controls (66.3 &#177; 13.3% and 63.4 &#177; 13.8% vs. 73.1 &#177; 2.8% and 72 &#177; 3.2%). SctO2 at rest is significantly linearly correlated with peak VO2 (r = 0.602), oxygen uptake efficiency slope (r = 0.501), and brain natriuretic peptide (r = -0.492), all of which are recognized prognostic and disease severity markers, indicating its potential prognostic value. SctO2 is determined mainly by end-tidal CO2 pressure, mean arterial pressure, and hemoglobin in the HF population. This article demonstrates a protocol that integrates SctO2 using NIRS into incremental CPET on a calibrated bicycle ergometer.

This protocol integrated near-infrared spectroscopy into conventional cardiopulmonary exercise testing to identify the involvement of the cerebral hemodynamic response in exercise intolerance in patients with heart failure.

Cerebral hypo-oxygenation during rest or exercise negatively impacts the exercise capacity of patients with heart failure with reduced ejection fraction ...

Phenotyping Mouse Pulmonary Function In Vivo with the Lung Diffusing Capacity

The mouse is now the primary animal used to model a variety of lung diseases. To study the mechanisms that underlie such pathologies, phenotypic methods are needed that can quantify the pathologic changes. Furthermore, to provide translational relevance to the mouse models, such measurements should be tests that can easily be done in both humans and mice. Unfortunately, in the present literature few phenotypic measurements of lung function have direct application to humans. One exception is the diffusing capacity for carbon monoxide, which is a measurement that is routinely done in humans. In the present report, we describe a means to quickly and simply measure this diffusing capacity in mice. The procedure involves brief lung inflation with tracer gases in an anesthetized mouse, followed by a 1 min gas analysis time. We have tested the ability of this method to detect several lung pathologies, including emphysema, fibrosis, acute lung injury, and influenza and fungal lung infections, as well as monitoring lung maturation in young pups. Results show significant decreases in all the lung pathologies, as well as an increase in the diffusing capacity with lung maturation. This measurement of lung diffusing capacity thus provides a pulmonary function test that has broad application with its ability to detect phenotypic structural changes with most of the existing pathologic lung models.

Phenotyping Mouse Pulmonary Function In Vivo with the Lung Diffusing Capacity

We describe a means to quickly and simply measure the lung diffusing capacity in mice and show that it is sufficiently sensitive to phenotype changes in multiple common lung pathologies. This metric thus brings direct translational relevance to the mouse models, since diffusing capacity is also easily measured in humans.

The mouse is now the primary animal used to model a variety of lung diseases. To study the mechanisms that underlie such pathologies, phenotypic methods ...

Biology

Assessing Lung Function with Hyperpolarized Xenon-129 MRI

Quantitative Measure of Lung Structure and Function Obtained from Hyperpolarized Xenon Spectroscopy

Hyperpolarized Xenon-129 (HXe) magnetic resonance imaging (MRI) provides tools for obtaining 2- or 3-dimensional maps of lung ventilation patterns, gas diffusion, Xenon uptake by lung parenchyma, and other lung function metrics. However, by trading spatial for temporal resolution, it also enables tracing of pulmonary Xenon gas exchange on a ms timescale. This article describes one such technique, chemical shift saturation recovery (CSSR) MR spectroscopy. It illustrates how it can be used to assess capillary blood volume, septal wall thickness, and the surface-to-volume ratio in the alveoli. The flip angle of the applied radiofrequency pulses (RF) was carefully calibrated. Single-dose breath-hold and multi-dose free-breathing protocols were employed for administering the gas to the subject. Once the inhaled Xenon gas reached the alveoli, a series of 90&#176; RF pulses was applied to ensure maximum saturation of the accumulated Xenon magnetization in the lung parenchyma. Following a variable delay time, spectra were acquired to quantify the regrowth of the Xenon signal due to gas exchange between the alveolar gas volume and the tissue compartments of the lung. These spectra were then analyzed by fitting complex pseudo-Voigt functions to the three dominant peaks. Finally, the delay time-dependent peak amplitudes were fitted to a one-dimensional analytical gas-exchange model to extract physiological parameters.

Author Spotlight: Advancing Lung Disease Research with Free-Breathing Hyperpolarized Xenon-129 MRI

The manuscript presents a detailed protocol for using hyperpolarized Xenon-129 chemical shift saturation recovery (CSSR) to trace pulmonary gas exchange, assess the apparent alveolar septal wall thickness, and measure the surface-to-volume ratio. The method has the potential to diagnose and monitor lung diseases.

Hyperpolarized Xenon-129 (HXe) magnetic resonance imaging (MRI) provides tools for obtaining 2- or 3-dimensional maps of lung ventilation patterns, gas ...

Dual Test Gas Pulmonary Diffusing Capacity Measurement During Exercise in Humans Using the Single-Breath Method

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