Nasal Nitric Oxide Measurement in Primary Ciliary Dyskinesia. A Technical Paper on Standardized Testing Protocols

Abstract

Nasal nitric oxide concentrations are extremely low in primary ciliary dyskinesia (PCD), and measurement of this nasal gas is recommended as a PCD diagnostic test in cooperative patients aged 5 years and older. However, nasal nitric oxide measurements must be performed with chemiluminescence analyzers using a standardized protocol to ensure proper results, because nasal nitric oxide values can be influenced by various internal and external factors. Repeat nasal nitric oxide testing on separate visits is required to ensure that low diagnostic values are persistent and consistent with PCD. This technical paper presents the standard operating procedures for nasal nitric oxide measurement used by the PCD Foundation Clinical and Research Centers Network at various specialty centers across North America. Adherence to this document ensures reliable nasal nitric oxide testing and high diagnostic accuracy when employed in a population with appropriate clinical phenotypes for PCD.

Keywords: Kartagener syndrome; nasal nitric oxide; primary ciliary dyskinesia.

Figure 1.

Nasal nitric oxide (NO) concentrations in subjects with primary ciliary dyskinesia (PCD; 17–180 parts per billion [ppb]) compared with healthy control subjects (543–976 ppb) and subjects with cystic fibrosis (CF; 241–896 ppb), sinusitis (862–3,601 ppb), Young’s syndrome (330–1,532 ppb), and bronchiectasis (516–1,098 ppb). Though nasal NO concentration in parts per billion is shown on the y-axis, one should always use nasal NO production values in nanoliters per minute. Because the NO analyzer used to perform these measurements has a flow sampling rate of 0.25 L/min, values below 308 ppb are equivalent to the accepted nasal NO cutoff of 77 nl/min. Only nasal NO values in patients with cystic fibrosis routinely fall below this cutoff value. ^#P < 0.0001. Reprinted by permission from Reference .

Figure 2.

(A) Hierarchical summary receiver operating characteristic curve (HSROC) for 12 evaluated studies in a detailed meta-analysis of nasal nitric oxide (nNO) values for diagnosing primary ciliary dyskinesia (PCD) in patients with classic ciliary electron microscopy defects and/or disease-causing variants in a known PCD gene. The summary sensitivity for nNO testing (red square) is 97.6% (range 92.7–99.2) and specificity is 96.0% (range 87.9– 98.7). Reprinted from Reference . (B) Scatterplot of nNO values (linear scale; nl/min) versus age for University of North Carolina disease control subjects and a subset of subjects with PCD with normal electron micrographs but biallelic mutations in DNAH11 (solid circles). Individual nNO measurements are shown for disease control subjects: asthma (open circles), cystic fibrosis (open squares), and chronic obstructive pulmonary disease (solid triangles). The dashed line represents the PCD cutoff nNO value of 77 nl/min. Reprinted from Reference . (C) Validation nNO data from six PCD specialty sites in the Genetic Disorders of Mucociliary Clearance Consortium. Confirmed cases of PCD had a classic electron microscopy ultrastructural defect and/or biallelic mutations in one known PCD gene. nNO in 155 consecutive individuals investigated for possible PCD was below the 77 nl/min cutoff (dashed line) in 70 of 71 confirmed PCD cases. The open circles represent individuals with PCD confirmed by electron microscopy ultrastructural defect. The solid circles represent individuals with PCD confirmed by genetic testing alone. Reprinted from Reference .

Figure 3.

(A) Nasal catheters with surrounding nasal sponge olives in three different sizes. Note the built-in air filter at the opposite end in each of these disposable catheters. The neonatal-size catheter is not shown here. Manufactured by DirectMed. (B) Disposable cardboard tube resistor. Patients securely seal their lips around the open end and gently blow in a sustained and prolonged manner into this resistor to achieve velum closure and to prevent dilution of sampled nasal gas with lower airway flow. Note the pinhole opening (white arrowhead) in the red resistor cap, which provides approximately 3–5 cm H₂O of airway resistance. Manufactured by DirectMed. (C) A 3-year-old child performing an exhalation against resistance nasal nitric oxide measurement using a child’s party favor resistor. Note the tape occluding the distal end of the party favor and the sustained exhalation maneuver keeping the party favor in an extended position long enough to achieve a plateau value. (D) A 7-year-old child performing an exhalation against resistance nasal nitric oxide measurement using a cardboard tube mouth resistor. The lips are securely tightened around the resistor to avoid leaking.

Figure 4.

Ambient nitric oxide (NO) washout technique. When the ambient NO value is greater than 10 ppb for a tidal breathing test or greater than 50 ppb for a resistor breathing test, an NO washout should be performed before and during the test. The technician places the nasal catheter into the desired naris and, using the NO scrubber device, allows NO-free air to blow into the nonoccluded nostril for 1 minute as the patient continues to breathe via their nonoccluded nostril. This source of NO-free air is continually blown into the nonoccluded nostril until testing is complete.

Figure 5.

Nasal nitric oxide (NO) curve of a patient exhaling into a cardboard resistor. Note that the first plateau may seem acceptable, but the following two plateaus are more than 10% higher, reflecting improved exhalation technique upon repeat maneuvers. Thus, the last two plateau values would be used to calculate the nasal NO in this case, whereas the first plateau would be discarded. ppb = parts per billion.

Figure 6.

Nasal nitric oxide (NO) curve of a cooperative 4-year-old child performing nasal NO measurement with a tidal breathing technique. Note the reproducibility and stability of numerous tidal peaks just above 140 parts per billion (ppb; white arrows).

Figure 7.

Nasal nitric oxide (NO) tracing of a 3-year-old child. For the first 40 seconds, the reproducible and maximal tidal peaks fall near 250 parts per billion (ppb; black arrows). However, after 60 seconds, the maximum reproducible peaks are higher and fall near 325–350 ppb (white arrows). Thus, tidal breathing tests should record for at least 60 seconds to ensure that reproducible, maximal tidal peaks are detected. One peak is clearly an outlier (gray arrow) because it is not reproducible and more than 10% above the other tidal peaks.

Figure 8.

Nasal nitric oxide (NO) tracing of a 3-year-old child performing a tidal breathing maneuver. Although there is a reproducible set of tidal peaks near 250 parts per billion (ppb; black arrows), there are higher reproducible tidal peaks near 400 ppb (white arrows). These higher peaks (white arrows) should be chosen as the maximal values to use for final nasal NO calculations.

Figure 9.

Nasal nitric oxide (NO) tracing from an uncooperative, crying child performing a tidal breathing maneuver. With spontaneous, prolonged exhalation maneuvers during crying, this child produces plateau values that can also be used as tidal peaks (peak numbers 2 and 4). ppb = parts per billion.