Investigations in Nasal Disease
Summary
The investigation of upper airway disorders is important because such problems can have a significant impact on patients’ quality of life; this can be ameliorated by accurate therapy. Presentation of severe disorders with significant morbidity and even mortality (e g., Wegener granulomatosis) can occur in the upper airways, and early diagnosis and effective management can prevent serious consequences. In addition, upper respiratory tract problems exacerbate those in the lower respiratory tract and may extend to involve it.
Allergists, respiratory physicians, and pediatricians, as well as ear, nose, and throat (ENT) specialists, need to be able to recognize and diagnose nasal disease accurately. This chapter covers tests for allergy, nasal sampling, the evaluation of nasal patency, microbiology, mucociliary clearance investigation, and various blood tests that are useful in the evaluation of rhinosinusitis.
Note
Upper respiratory tract problems may exacerbate those in the lower respiratory tract, and vice versa.
Introduction
Nasal disease has many different diagnoses that cannot be differentiated merely by history taking and physical examination. Additional diagnostic tests are often needed for accurate investigation of upper airway disorders. Some disorders of the upper respiratory tract are severe, with significant morbidity and even mortality; therefore, immediate and accurate diagnosis and effective management are essential. In other cases, there can be a significant impact on patients’ quality of life, which calls for well-directed treatment. Another important factor is the united airway concept: upper respiratory tract problems are an independent risk factor for developing lower respiratory tract disorders. Furthermore, sinonasal disease can be the cause of exacerbation of lower respiratory symptoms. Early diagnosis gives the opportunity for prevention of severe complications. This united airway concept is true not only for allergic and nonallergic rhinitis, which can progress to asthma, but also for rhinosinusitis, which can be the first presentation of Wegener granulomatosis and Churg-Strauss syndrome. The nose is the first site of allergen, microbial, and particle deposition, which is reflected in its highly developed local innate and adaptive immune system. Defects in mucociliary clearance can lead to severe problems, as can be seen in primary ciliary dyskinesia (PCD) and cystic fibrosis (CF). Finally, the nose is an easily accessible area for investigation of mechanisms of disease and therapy. This chapter offers a basic introduction to methods used in rhinology, including their applicability, specificity, and sensitivity.
Note
Upper airway disorders can have a significant impact on patients’ quality of life.
Caution
Sinus disorders can lead to significant morbidity and even mortality, which can be prevented with early diagnosis and effective management.
Tips and Tricks
In cases of chronic rhinosinusitis, consider additional diagnostic tests for systemic pathology, such as cystic fibrosis, Wegener granulomatosis, and immunodeficiency.
Allergy Tests
Allergen-specific immunoglobulin E (IgE) is the triggering factor of symptoms and of the underlying inflammatory process of allergic rhinitis. Diagnostic tests aim to demonstrate the presence and/or functional relevance of such IgE. The presence of specific IgE alone (sensitization) does not necessarily imply clinical relevance, and all results need interpreting in light of the patient′s history.1
The presence of specific IgE can be demonstrated either in vivo (skin prick tests) or in vitro by detecting serum allergen-specific IgE (radioallergosorbent test [RAST], ImmunoCAP [CAP-RAST, Pharmacia Diagnostics AB, Uppsala, Sweden]). Skin prick tests are considered the first-line approach for the detection of allergic sensitization, due to their speed, efficiency, safety, and relatively low cost. The biological assays (CAP-RAST) are similarly sensitive, but they are more expensive and time-consuming, require laboratory facilities, and give delayed results.
The functional role of specific IgE can be demonstrated in vitro by basophil activation tests or in vivo by allergenspecific provocation. Basophil degranulation tests require a special laboratory apparatus and are not routinely used.
Nasal allergen provocation tests involve delivery of likely allergens to the nose with monitoring of the response, aiming to reproduce the reaction occurring during natural allergen exposure. There are a variety of substances, measurements, and evaluation techniques for nasal challenges ( Fig. 6.1 ). Allergen challenges allow demonstration of the causal role of the allergen. The same happens with occupational substances or with aspirin, although in these cases the mechanism may be non IgE-mediated. Non-allergen-specific challenges evoke a nonspecific inflammatory response and demonstrate the existence of nasal hyperreactivity.
Skin Prick Test
The skin prick technique is the gold standard method, and tests are highly reproducible with a trained investigator.2 They should be performed according to a rigorous methodology, with standardized diagnostic extracts, and always must include a negative (saline diluent) and a positive (histamine hydrochloric acid [HCl] 0.1%) control. They should be read at the peak of reaction (at ~15 minutes) by measuring the dimension of wheals ( Fig. 6.2 ). The diagnostic significance of late reactions is not known. The scoring of the positivity is given according to recommendations of the European Academy of Allergy and Clinical Immunology (EAACI), and the interpretation of a positive test must be integrated with the clinical history, as a positive skin prick test does not always imply a clinically relevant sensitization.
A false-positive reaction may occur if dermagraphism is present, but this can be ruled out with the use of a negative control. False-negative reactions may occur, most commonly as a result of interfering drugs, such as systemic antihistamines. Because of this, antihistamines must be discontinued at least 5 days before testing. Tricyclic anti-depressants can interfere with test results and should be stopped at least 3 weeks before testing. Topical corticosteroids used on the skin site tested and high-dose oral corticosteroids may also decrease reactions. Other causes of false-negative skin prick tests are weak potency of the extract and improper technique. Antileukotrienes do not interfere with skin prick tests and can be continued.
Intradermal tests are not used for the diagnosis of respiratory allergy, as they do not perform better than skin prick tests and can induce false-positive results. On the other hand, intradermal tests remain an essential part of the diagnostic work-up for hymenoptera and drug allergy. The scratch test is no longer in use.
Atopy patch tests involve epicutaneous patch tests with allergens known to elicit IgE-mediated reactions. Commercial reagents are available for a few allergens and have been standardized regarding the use of vehicle and dose–response relationships. A subset of patients with atopic dermatitis show only atopy patch test positivity, while specific IgE to the same allergen remains negative,3 but the atopy patch test is usually not relevant for the diagnosis of respiratory allergies.
Serum Total IgE
Serum total IgE can be measured using either radioimmunoassay or enzyme assay. Levels of IgE increase from birth to adolescence, then decrease to reach a plateau after the age of 20 to 30 years in normal subjects. In adults, levels > 100 to 150 kU/L are considered to be elevated. Total IgE may be increased in allergy but is not always elevated in rhinitis; it is also elevated in smokers and in parasitic diseases. Thus, because an increase of total IgE correlates weakly with the presence of allergic diseases, the measurement of serum total IgE should no longer be used for screening or allergy diagnosis.4
Detection of Allergen-specific IgE in Serum
The RAST, first used for the measurement of serum allergen-specific IgE, has been replaced by immune-enzymatic methods, including the widely used CAP-RAST. With these assays, the level of specific IgE is expressed as kU/L, according to calibration curves, and the cut-off IgE level above which the test is positive is usually 0.35 kU/L. However, some sensitized subjects have an IgE level below this cut-off. The measurement of serum-specific IgE is usually less sensitive than skin prick tests,5 and the worst correlations between skin prick tests and IgE assays are obtained with mold, food extracts, and nonstandardized extracts. In general, the correlation between a strongly positive response to a skin prick test and the detection of serum-specific IgE and between a negative response to a skin prick test and the lack of detection of serum-specific IgE is very good. As in skin prick tests, the presence or absence of specific IgE in the serum does not necessarily indicate a clinically relevant allergy. As mentioned above, for inhalant allergens, skin prick test responses represent the first-line diagnostic methods, and when they correlate with the clinical history, in vitro tests are not necessary.4
It has been recently shown that in a subset of patients, the presence of specific IgE in the nasal mucosa can be demonstrated when skin prick tests are negative (entopy).6 Nonetheless, the measurement of IgE in nasal secretions is not a routine assay, and such patients could be identified by nasal allergen challenge.
The presence of functionally relevant specific IgE can be demonstrated by putting the allergen into contact with basophils and subsequently detecting their activation by cytofluorometry. This basophil degranulation test has been proposed for specific conditions, such as drug allergies, but is not recommended for respiratory allergy diagnosis and is used only for research purposes.
Nasal Provocation Tests
Nasal Allergen Challenge
This procedure, also known as the specific nasal provocation test (SNPT) or nasal allergen-specific nasal challenge (ASNC), involves the delivery of a small quantity of the allergen into one or preferably both nostrils to elicit an allergic reaction, if allergen-specific IgE is present in the nasal mucosa. By using progressively increasing amounts (or concentrations) of the allergen, a threshold dose can be established. Recently, the availability of recombinant purified allergens has suggested the possibility of performing challenges with each specific allergenic protein, but the role of such an approach in clinical practice is still not defined.
The main advantages of NAC are simplicity of execution, low cost, and safety. The main disadvantage is that only one allergen can be tested at a time. The procedure for nasal allergen provocation has been rationalized;7 however, it is still poorly standardized, and the technical details (amount of allergen, interval between doses, dilutions, and positivity criteria) vary among practices. The main indications for NAC are to diagnose allergic rhinitis in patients with negative or doubtful skin prick tests, to identify the clinically relevant allergen(s) in polysensitized subjects, to study the allergic reaction, and to evaluate the effects of treatment.
Contraindications to NAC are acute bacterial or viral rhinosinusitis, acute exacerbation of allergic disease, history of previous anaphylactic reaction, severe systemic diseases, pregnancy, and recent (within the past 6 to 8 weeks) ENT surgery.
The general procedure is as follows: (1) Allow the patient to adapt to the challenge room for > 15 minutes, then inspect the nasal cavity. (2) Spray or apply saline as a negative control. (3) Perform measurement(s). (4) Instruct the patient to avoid nasal breathing. (5) Apply the allergen into one or both nostrils and reevaluate after 10 to 15 minutes. (6) Proceed with increasing concentrations until a reaction occurs or the top dose is reached.
When an allergen is introduced into the nose, the IgE-mediated reaction immediately takes place, and the classic symptoms appear in minutes. Symptoms usually subside within 4 to 6 hours, but they may persist or reappear as a late-phase reaction.
Causes of a false-negative reaction include weak extract and counteracting medication, such as nasal anti-histamine (1-day withdrawal), oral antihistamine (3-day withdrawal), or nasal steroid (7-day withdrawal).
False-positive reactions can occur due to the nasal cycle (which is the reason for bilateral challenge) or to nasal hyperreactivity with a nonspecific response (which is the reason for an initial placebo challenge).
NAC can be evaluated in many different ways. The most common is the measurement of the four classic nasal symptoms by an ordinal scale (0 = absent to 3 = severe), the test being positive if an increase in score of 5 or more points is obtained. Alternatively, a visual analogue scale can be used. Also, sneezes can be counted and nasal secretions weighed.
Quantitative assessment of the NAC can be made by measuring nasal flow or resistance by nasal peak flow meter, acoustic rhinometry, or rhinomanometry (vide infra).
Other possible modalities to evaluate the effect of allergen challenge are the assessment of the inflammatory infiltrate by nasal scraping/brushing (with differential cell count) and the measurement of specific mediators in nasal lavage, including tryptase, α2-microglobulin, albumin, leukotrienes, interleukins, and eosinophil cationic protein.
Nasal Challenge with Lysine Aspirin
Nasal challenge with lysine aspirin is not an allergen challenge specifically, as no IgE-mediated mechanism is involved. Nevertheless, it has the value of a specific challenge and is used to diagnose aspirin intolerance in the context of aspirin hypersensitivity with respiratory manifestations. The nasal aspirin challenge was introduced later than the oral and bronchial challenges but has gained popularity because it rarely induces systemic reactions. Nasal aspirin challenge is possible in patients with severe asthma in whom oral and bronchial aspirin challenges are contraindicated.
The aspirin challenge is sufficiently standardized and reproducible,8 although the possibility of false-negative results exists, and the negative predictive value is lower than for oral and bronchial challenges. For this reason, it is agreed that where aspirin intolerance is suspected and the nasal challenge is negative, an oral challenge must be performed. The nasal aspirin challenge must be performed under medical supervision. Oral corticosteroids, nasal steroids, and antileukotrienes should be discontinued at least 7 days before, whereas the withdrawal period is 3 days for antihistamines and 24 hours for decongestants and cromones. Lysine-aspirin solutions at 0.1, 1, and 2 M are used at 45-minute interval steps. The evaluation of the result includes clinical symptoms and instrumental measurements (acoustic rhinometry, anterior rhinomanometry). Pulmonary function must be monitored during the challenge.
Simpler and safer alternatives to the aspirin provocation test are highly desirable. The cellular antigen stimulation test (CAST) has been proposed as an alternative, but a recent study using CAST to measure cysteinyl leukotrienes (cys-LTs) before and after the challenge showed that although the leukocytes of patients with aspirin sensitivity produce higher amounts of cys-LTs, the assay had a sensitivity of 25%, a specificity of 92.3%, and positive and negative predictive values of 28.7% and 90.7%, respectively. The low sensitivity and predictive values limit the clinical usefulness of this test.9
Nonspecific Nasal Challenges
Nasal hyperreactivity occurs when the nasal mucosa responds with symptoms to nonspecific stimuli that do not cause a reaction in normal subjects. It is common in patients with allergic or other inflammatory forms of rhinitis.10 A variety of stimuli can be used to evoke nasal hyperreactivity. These may directly act on a single receptor (e.g., histamine, adenosine monophosphate (AMP), and methacholine) or activate a more complex mechanism (e.g., mannitol, capsaicin, hyperosmolar solutions, and cold air).
The results obtained are often difficult to interpret due to the heterogeneity of methods, doses, and outcomes. For example, histamine and methacholine are both able to evoke more pronounced nasal reaction in subjects with rhinitis than in healthy controls,11 but only histamine is able to modify the nasal resistance. Cold, dry air is capable of differentiating between patients with perennial nonallergic rhinitis and healthy subjects, but histamine is not.12 In addition, mannitol nasal challenge seems unable to activate mast cells, although a dose–response relationship in eliciting symptoms has been reported for this test.
The AMP challenge is relatively simple and reproducible and more sensitive than the histamine challenge.13 It is thought to predict the response to nasal steroids and acts as a surrogate marker to evaluate the antiinflammatory effects of drugs.
The nasal capsaicin challenge displays a dose-dependent response14 and is able to detect nasal hyperreactivity in seasonal allergic rhinitis. Because of its selectivity for sensory nerves, it is mainly used for experimental purposes to study cough reflexes.
Nasal provocation with cold, dry air requires special apparatus and is currently used only for research purposes. Interestingly, cold air provocation has been reported to be able to discriminate between rhinitis alone and rhinitis associated with asthma, as well as between patients with nonallergic rhinitis and healthy subjects.12 Unlike asthma, a threshold dose discriminating between healthy and rhinitis subjects has not been unequivocally established.
Nasal Sampling
In sinonasal disease, it may be necessary to assess the level of inflammation in the nasal mucosa, as a diagnostic tool distinguishing inflammatory from neurogenic rhinitis and to evaluate the effect of therapy. A variety of approaches can be used; these include nasal lavage, nasal cytology, nasal biopsy, and nasal nitric oxide (NO) measurements. Each of these sampling techniques has its own strengths and weaknesses. Which technique is used depends on the diagnostic or research question asked.
A comparison of the different techniques is shown in Table 6.1.
Nasal Secretions
Nasal secretions contain inflammatory cells and mediators, reflecting nasal mucosal inflammation. They can be collected in several ways: blown secretions, nasal lavage, absorption in sinus packs or filter paper, or by microsuction.
In the first method secretions in the nasal airways are blown onto wax paper or a plastic wrap and then placed onto a glass slide. Microscopic evaluation allows the discrimination of epithelial cells from granulocytes. This method is simple, but gives little information about the nasal mucosa.
Nasal lavage is simple and quick, usually well tolerated, and provides a sample for evaluation of the secretion in the nasal lumen for proteins, cells, mediators, and cytokines. It involves the introduction of fluid into the nasal cavity and its recovery after a predetermined time. A range of techniques has been used to instill and recover fluid from the nasal cavity. Usually a volume of 2.5 to 5.0 mL 0.9% sodium chloride [NaCl], prewarmed to 37°C (98.6°F), is instilled within each nostril with an 80% recovery (range 65–90%). An agent to disrupt the disulphide bonds of the mucus polypeptide chains, such as Mesna (sodium 2-mercaptoethane sulphonate), can be included. In situations of extreme nasal blockage, the obstruction of the nasal lumen will limit the amount of fluid that can be retained within the nasal cavity, and smaller lavage volumes need to be used. The consistency of the findings in allergic and infective rhinitis for a range of different measures in nasal lavage fluid supports the concept that this method of nasal evaluation provides reliable and relevant information. However, normalization of recovery of the fluid can be difficult. Another disadvantage is that initial histamine levels are high, probably reflecting bacterial origin, and initial washouts are needed before histamine levels can be reliably measured. Also, there is considerable variability between subjects in luminal eosinophil recruitment and activation.
Sinus packs or filter paper can be used to collect nasal secretions without diluting the cells and mediators. In this method, preweighed sinus packs are placed on the floor of the nasal cavity between the septum and inferior turbinate for 5 minutes and are then placed in a Falcon tube15 and washed with 3 mL of 0.9% NaCl solution. The sinus pack is then placed into the shaft of a syringe and squeezed by the piston. After this first pressure, the shaft containing the sinus pack is placed into a Falcon tube and centrifuged at 1500 g for 10 minutes to recover all fluid. If irritation of the nasal mucosa is an issue, thin filter paper that can be inserted without touching the nasal mucosa can be substituted for sinus packs. The amount of secretion that can be absorbed in this way is more limited.
Nasal secretions can also be collected by direct aspiration (the microsuction technique). The samples are collected by repeated aspiration into a preweighed plastic sampling tube immediately followed by aspiration of a known volume (1.0 mL) of phosphate-buffered saline (PBS) containing 10% Mesna, which disrupts disulphide bonds of the mucus polypeptide chains. The direct aspiration system combines the advantages of minimal irritation of the nasal mucosa with the facility to determine concentrations per gram of secretion.16
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