Gosepath et al, 2005

Rondón C et al, 2007

Kim & Jang, 2011

Pirila & Nuutinen, 1998

Wihl, 1986

In another study conducted by our group, 21 allergic patients were submitted to *Dermatophagoides pteronyssinus* specific NPT (BialAristegui, Bilbao, Spain). Secretions, namely tears and nasal secretions, were collected after NPT and inflammatory mediators, such as interleukins and chemokines were measured (data not published). These inflammatory mediators were measurable only in 21% of the tear samples and in 71.5% of the nasal secretion samples. According to these findings, nasal secretions recovery could be acceptable to be considered as an objective tool in the evaluation of inflammatory mediators. However we could not find out a pattern of mediator release since the inflammatory mediators were inconsistently detected in the different samples.

Although the analysis of immunological parameters has been described as an objective approach to monitor the response to NPT, in our experience, these laboratorial measurements are difficult to perform because of the scheduling of sampling. Additionally, the cost-effectiveness of these procedures does not allow its implementation in the clinical practice. It should be reserved to investigational research.

#### **3.2.4 Assessment on nasal Nitric Oxide**

Determination of nasal Nitric Oxide (nNO) provides an indirect measure of the inflammation of the nasal mucosa. A decrease in nNO levels with NPT coincided with maximal symptom intensity, in five patients with pollen-induced allergic rhinitis (Kharitonov et al, 1997). Although nNO promises as a diagnostic non-invasive management tool, its value in nasal pathology is still not clear, mainly due to the lack of standardization of the test. Different methods of measurement have been used in published studies and the results reported are not comparable (Dordal et al, 2011).

#### **3.3 Criteria for positive NPT**

Besides diverse combined criteria have been discussed in the literature none of them are standardized criteria to define NPT positivity. This is summarized in the Table 2.

#### **4. NPT as a diagnostic approach in respiratory allergy**

The first allergen provocation test was performed in 1873, by Blackley, who elicited a nasal response after an application of fresh pollen to the membrane of his own nostrils (Blackley, 1873). Currently, the indications of NPT are widely known. In this manuscript, the applicability of NPT as a diagnostic tool of allergic rhinitis was discussed. But the usefulness of NPT is not restricted to the diagnostic approach of allergic rhinitis. Supported by the concept of *"one airway, one disease",* several studies have pointed out that the NPT is a good alternative to Bronchial Provocation Test (BPT), even in the absence of nasal symptoms. In spite of BPT being a standardized diagnostic tool, it is not frequently used in clinical practice because of its technically and methodologically requirement. Indeed, NPT is safer and better tolerated

In another study conducted by our group, 21 allergic patients were submitted to *Dermatophagoides pteronyssinus* specific NPT (BialAristegui, Bilbao, Spain). Secretions, namely tears and nasal secretions, were collected after NPT and inflammatory mediators, such as interleukins and chemokines were measured (data not published). These inflammatory mediators were measurable only in 21% of the tear samples and in 71.5% of the nasal secretion samples. According to these findings, nasal secretions recovery could be acceptable to be considered as an objective tool in the evaluation of inflammatory mediators. However we could not find out a pattern of mediator release since the inflammatory

Although the analysis of immunological parameters has been described as an objective approach to monitor the response to NPT, in our experience, these laboratorial measurements are difficult to perform because of the scheduling of sampling. Additionally, the cost-effectiveness of these procedures does not allow its implementation in the clinical

Determination of nasal Nitric Oxide (nNO) provides an indirect measure of the inflammation of the nasal mucosa. A decrease in nNO levels with NPT coincided with maximal symptom intensity, in five patients with pollen-induced allergic rhinitis (Kharitonov et al, 1997). Although nNO promises as a diagnostic non-invasive management tool, its value in nasal pathology is still not clear, mainly due to the lack of standardization of the test. Different methods of measurement have been used in published studies and the

Besides diverse combined criteria have been discussed in the literature none of them are

The first allergen provocation test was performed in 1873, by Blackley, who elicited a nasal response after an application of fresh pollen to the membrane of his own nostrils (Blackley, 1873). Currently, the indications of NPT are widely known. In this manuscript, the applicability of NPT as a diagnostic tool of allergic rhinitis was discussed. But the usefulness of NPT is not restricted to the diagnostic approach of allergic rhinitis. Supported by the concept of *"one airway, one disease",* several studies have pointed out that the NPT is a good alternative to Bronchial Provocation Test (BPT), even in the absence of nasal symptoms. In spite of BPT being a standardized diagnostic tool, it is not frequently used in clinical practice because of its technically and methodologically requirement. Indeed, NPT is safer and better tolerated

Lebel B et al, 1988 Symptom score Lebel Symptom Score Scale: Positive if <sup>≥</sup>

response Description of positivity criteria

Δ ≥ 4, considering Δ = (obstruction score

5 (maximum possible score 11 points)

+ rhinorrhea score) after NPT - (obstruction score + rhinorrhea score)

standardized criteria to define NPT positivity. This is summarized in the Table 2.

mediators were inconsistently detected in the different samples.

practice. It should be reserved to investigational research.

results reported are not comparable (Dordal et al, 2011).

Reference Assessment of nasal

Hytonen et al, 1997 Symptom score

**4. NPT as a diagnostic approach in respiratory allergy**

**3.2.4 Assessment on nasal Nitric Oxide**

**3.3 Criteria for positive NPT**

Nasal Provocation Test in the Diagnosis of Allergic Rhinitis 165

is produced after local allergic reactions (Togias, 2003). The link between local exposure to allergen and distant response has been clarified. Although some authors defend that this systemic response could result from allergen entering in the systemic circulation from the local of exposure (Hens et al, 2007) this could activate circulating basophils, inducing an anaphylactic reaction, which is a rare condition (Togias, 2004). Both systemic cell circulation and the nervous system activation are two major ways through which local allergic reactions propagate. Mast cell mediators locally released, increase the expression of adhesion molecules on postcapillary venules. This can lead to homing of circulating leukocytes, which may infiltrate distant tissues. This cell recirculation and focalization makes the IgE mediated allergic disease a dynamic and systemic process. Pereira C showed that this cell response starts at an early stage, in parallel with the immediate allergic response (Pereira, 2009). The IgE mediated response induces immunolymphatic involvement of the adjacent structures. This amplifies the allergic response to loco-regional lymphoid organs, while circulating leukocytes recirculation compromises the primary lymphoid organs (thymus and bone marrow). These central organs are responsible for the systemic immune response induced by a localized allergen challenge, in this case, a nasal challenge (Pereira, 2009). The nervous system activation could be involved by, any or both pathways, namely neurogenic inflammation and neuronal reflexes. Neurogenic inflammation is characterized by specific neuromediators closely related to neuro-immune-endocrine system, and it is both a stimulus to and a consequence of allergic inflammation. The naso-nasal and the naso-ocular reflexes are some examples of the role of the nervous system in the propagation of the allergic disease. They seem to be predominantly mediated by parasympathetic and cholinergic pathways, respectively (Baroody et al, 1994, 2008). Histamine release during the acute response to allergen and substance P seem to have an important role in these neural mechanisms (Baroody et al, 1994, 2008; Fujishima et al 1997; Micera et al, 2008; O'Meara et al, 2005; Sheahan et al, 2005). Multiple evidences support a close interaction and influence of the nose on contiguous and distant organs via neural reflex and systemic inflammatory processes (Baroody, 2011). In summary, a local triggered allergenic inflammation is systematically extended, with the early connection of the immune central organs. Independently of the involved pathway, immediate symptoms are clinically

Besides the limitations of NPT, this is a feasible and easily method to be performed, since the nasal cavities provide easy access to specific provocation. The concept of "*One airway, one disease*" allows assuming the similarity of response to the provocation of both the upper and lower airways, so the nasal allergic reaction could be accepted as predictor of bronchial response. Supported by the concept of the bidirectional "unified airway" respiratory inflammation, a local provocation test is useful in the diagnosis of allergic respiratory disease. Concerning these aspects, the NPT is the method of choice for the reproducibility of the allergic reaction (Litvyakova & Baraniuk, 2001; Loureiro, 2001; Mellilo, 1997; Naclerio & Norman, 1998). Thus the NPT may be considered a model of respiratory provocation test, easy to perform, in the demonstration of the immediate and late phase of type I

According to all the mentioned above, the clinical symptom score is widely used in clinical practice, alone or associated to objective measurements of nasal obstruction, namely nPIFR,

manifested.

hypersensitivity reaction.

**5. Characterization of NPT score symptom response**

method in asthmatic patients than BPT (Hervás et al, 2011; Marcucci *et al*, 2007; Oddera *et al*, 1998). So, NPT has been used to the diagnosis of asthma, as reviewed by Olive Pérez, 1997. Thus, based on the *united airways disease* concept, the NPT could be considered as a model of specific provocation test that is easy and quick to perform, in the demonstration of the immediate and late phase response of type I hypersensitivity reaction. It is well known that the nose is an integral part of the upper airway, and anatomically related to several airway structures, such as ears and paranasal sinuses, and as well the eyes. There is an epidemiological relationship between rhinitis and asthma. Rhinitis and asthma are often associated, rhinitis typically precedes the development of asthma and can contribute to insufficient asthma control (Compalati et al, 2010). On the other hand, in cross-sectional and longitudinal studies, the vast majority of patients with asthma have rhinitis, and rhinitis is a major independent risk factor for asthma (Togias, 2003). Treating allergic rhinitis would probably ameliorate other associated upper airway diseases such as acute rhinosinusitis, nasal polyposis, adenoidal hypertrophy, and OME (Marple, 2010). In addition to improve allergic rhinitis outcome, the treatment of subjacent inflammatory disorder reduces asthma-associated health care consuming. A close interaction between the nose and contiguous or distant organs was described and it has been progressively clarified, supporting this epidemiological and clinical relation (Baroody, 2011). The upper and lower airways are not anatomically and functionally distinct areas (Slavin, 2008). It is currently established that the impaired function of the upper airways causing nasal obstruction, retention of secretions, and disturbed conditioning of the inspired air plays an important role in the development of lower airway symptoms (Virchow, 2005). There are important relationships between both the nose and the paranasal sinuses and asthma. Apart from the intrinsic physiological interaction, extensive evidence exists to sustain the concept that the respiratory system functions as an integrated unit (Krouse, 2008), where rhinitis and asthma are manifestations of one syndrome, the chronic allergic respiratory syndrome, in both parts of the respiratory tract (Togias, 2003). It has been described that parallel immunopathological processes involve the upper airway generally occur in conjunction with lower airway diseases, and diffuse inflammation often affects mucosal surfaces of the middle ear, nose, sinuses, and tracheobronchial tree simultaneously (Krouse, 2008). Recent studies show that the deposition of allergen into the lower respiratory tract leads to increased inflammation of the upper respiratory tract, even if the patients are only suffering from allergic rhinitis (Virchow, 2005). Additionally, studies indicate that treatment of the upper respiratory tract inflammation reduces the manifestation of allergenassociated symptoms in the lower respiratory tract, and also have preventive effects if started early on the disease evolution (Bousquet & ARIA Workshop Group, 2001). Both asthma and allergic rhinitis have now been recognized as inflammatory diseases with similar manifestations in the mucous membranes of the upper (nose and paranasal sinuses) and lower respiratory tract (Virchow, 2005). There is increasing evidence that even in patients with rhinitis who do not have asthma, sub-clinical changes in the lower airways and inflammatory mediators can be detected (Compalati et al, 2010). These and other findings support that allergic diseases have a systemic component (Virchow, 2005). The interactive mechanisms of allergic rhinitis and associated conditions highlights the relevance of a bidirectional "unified airway" respiratory inflammation model. Currently, it is accepted that IgE mediated allergic reactions are not confined to the area where the trigger occurred, inducing a secondary systemic immune response (Braunstahl, 2005, 2006; Togias, 2004). The systemic inflammation

method in asthmatic patients than BPT (Hervás et al, 2011; Marcucci *et al*, 2007; Oddera *et al*, 1998). So, NPT has been used to the diagnosis of asthma, as reviewed by Olive Pérez, 1997. Thus, based on the *united airways disease* concept, the NPT could be considered as a model of specific provocation test that is easy and quick to perform, in the demonstration of the immediate and late phase response of type I hypersensitivity reaction. It is well known that the nose is an integral part of the upper airway, and anatomically related to several airway structures, such as ears and paranasal sinuses, and as well the eyes. There is an epidemiological relationship between rhinitis and asthma. Rhinitis and asthma are often associated, rhinitis typically precedes the development of asthma and can contribute to insufficient asthma control (Compalati et al, 2010). On the other hand, in cross-sectional and longitudinal studies, the vast majority of patients with asthma have rhinitis, and rhinitis is a major independent risk factor for asthma (Togias, 2003). Treating allergic rhinitis would probably ameliorate other associated upper airway diseases such as acute rhinosinusitis, nasal polyposis, adenoidal hypertrophy, and OME (Marple, 2010). In addition to improve allergic rhinitis outcome, the treatment of subjacent inflammatory disorder reduces asthma-associated health care consuming. A close interaction between the nose and contiguous or distant organs was described and it has been progressively clarified, supporting this epidemiological and clinical relation (Baroody, 2011). The upper and lower airways are not anatomically and functionally distinct areas (Slavin, 2008). It is currently established that the impaired function of the upper airways causing nasal obstruction, retention of secretions, and disturbed conditioning of the inspired air plays an important role in the development of lower airway symptoms (Virchow, 2005). There are important relationships between both the nose and the paranasal sinuses and asthma. Apart from the intrinsic physiological interaction, extensive evidence exists to sustain the concept that the respiratory system functions as an integrated unit (Krouse, 2008), where rhinitis and asthma are manifestations of one syndrome, the chronic allergic respiratory syndrome, in both parts of the respiratory tract (Togias, 2003). It has been described that parallel immunopathological processes involve the upper airway generally occur in conjunction with lower airway diseases, and diffuse inflammation often affects mucosal surfaces of the middle ear, nose, sinuses, and tracheobronchial tree simultaneously (Krouse, 2008). Recent studies show that the deposition of allergen into the lower respiratory tract leads to increased inflammation of the upper respiratory tract, even if the patients are only suffering from allergic rhinitis (Virchow, 2005). Additionally, studies indicate that treatment of the upper respiratory tract inflammation reduces the manifestation of allergenassociated symptoms in the lower respiratory tract, and also have preventive effects if started early on the disease evolution (Bousquet & ARIA Workshop Group, 2001). Both asthma and allergic rhinitis have now been recognized as inflammatory diseases with similar manifestations in the mucous membranes of the upper (nose and paranasal sinuses) and lower respiratory tract (Virchow, 2005). There is increasing evidence that even in patients with rhinitis who do not have asthma, sub-clinical changes in the lower airways and inflammatory mediators can be detected (Compalati et al, 2010). These and other findings support that allergic diseases have a systemic component (Virchow, 2005). The interactive mechanisms of allergic rhinitis and associated conditions highlights the relevance of a bidirectional "unified airway" respiratory inflammation model. Currently, it is accepted that IgE mediated allergic reactions are not confined to the area where the trigger occurred, inducing a secondary systemic immune response (Braunstahl, 2005, 2006; Togias, 2004). The systemic inflammation

is produced after local allergic reactions (Togias, 2003). The link between local exposure to allergen and distant response has been clarified. Although some authors defend that this systemic response could result from allergen entering in the systemic circulation from the local of exposure (Hens et al, 2007) this could activate circulating basophils, inducing an anaphylactic reaction, which is a rare condition (Togias, 2004). Both systemic cell circulation and the nervous system activation are two major ways through which local allergic reactions propagate. Mast cell mediators locally released, increase the expression of adhesion molecules on postcapillary venules. This can lead to homing of circulating leukocytes, which may infiltrate distant tissues. This cell recirculation and focalization makes the IgE mediated allergic disease a dynamic and systemic process. Pereira C showed that this cell response starts at an early stage, in parallel with the immediate allergic response (Pereira, 2009). The IgE mediated response induces immunolymphatic involvement of the adjacent structures. This amplifies the allergic response to loco-regional lymphoid organs, while circulating leukocytes recirculation compromises the primary lymphoid organs (thymus and bone marrow). These central organs are responsible for the systemic immune response induced by a localized allergen challenge, in this case, a nasal challenge (Pereira, 2009). The nervous system activation could be involved by, any or both pathways, namely neurogenic inflammation and neuronal reflexes. Neurogenic inflammation is characterized by specific neuromediators closely related to neuro-immune-endocrine system, and it is both a stimulus to and a consequence of allergic inflammation. The naso-nasal and the naso-ocular reflexes are some examples of the role of the nervous system in the propagation of the allergic disease. They seem to be predominantly mediated by parasympathetic and cholinergic pathways, respectively (Baroody et al, 1994, 2008). Histamine release during the acute response to allergen and substance P seem to have an important role in these neural mechanisms (Baroody et al, 1994, 2008; Fujishima et al 1997; Micera et al, 2008; O'Meara et al, 2005; Sheahan et al, 2005). Multiple evidences support a close interaction and influence of the nose on contiguous and distant organs via neural reflex and systemic inflammatory processes (Baroody, 2011). In summary, a local triggered allergenic inflammation is systematically extended, with the early connection of the immune central organs. Independently of the involved pathway, immediate symptoms are clinically manifested.

Besides the limitations of NPT, this is a feasible and easily method to be performed, since the nasal cavities provide easy access to specific provocation. The concept of "*One airway, one disease*" allows assuming the similarity of response to the provocation of both the upper and lower airways, so the nasal allergic reaction could be accepted as predictor of bronchial response. Supported by the concept of the bidirectional "unified airway" respiratory inflammation, a local provocation test is useful in the diagnosis of allergic respiratory disease. Concerning these aspects, the NPT is the method of choice for the reproducibility of the allergic reaction (Litvyakova & Baraniuk, 2001; Loureiro, 2001; Mellilo, 1997; Naclerio & Norman, 1998). Thus the NPT may be considered a model of respiratory provocation test, easy to perform, in the demonstration of the immediate and late phase of type I hypersensitivity reaction.

#### **5. Characterization of NPT score symptom response**

According to all the mentioned above, the clinical symptom score is widely used in clinical practice, alone or associated to objective measurements of nasal obstruction, namely nPIFR,

Nasal Provocation Test in the Diagnosis of Allergic Rhinitis 167

decline till 6 hours. Looking at the score of each nasal symptom (Figure 3), except for nasal obstruction, all of them followed the response pattern of total symptoms score, with a peak of symptoms at the fifth minute. Sneezing was the predominant symptom at the fifth minute, while nasal obstruction was the predominant symptom at the 30th minute and the sixth hour.

**5 minutes 30**

Fig. 3. Nasal symptom score after specific *Parietaria judaica* nasal provocation test (NPT): evolution of each nasal symptom (Legend: nasal congestion; pruritus; sneezing;

In another study mentioned above, the *Dermatophagoides pteronyssinus* specific NPT were performed in 34 children with OME. Those who had positive NPT, showed a response dynamics characterized by a rapid increase of symptoms score till a peak at the 5th minute (monitored till 1 hour), as shown in figure 4. Looking at the score of each nasal symptom (Figure 5), except for nasal pruritus, all followed the response pattern of total symptoms

**minutes**

**6 hours**

**0**

rhinorrea).

**before NPT**

score, with the peak of symptoms at the fifth minute.

> **before NPT**

(NPT); (Legend: total score; nasal score; non-nasal score).

**1st minute**

**5th minute**

Fig. 4. Score symptoms after *Dermatophagoides pteronyssinus* specific nasal provocation test

**30th minute** **1 hour**

**0.5**

**1**

**1.5**

**2**

**2.5**

rhinomanometry and acoustic rhinometry. The other methods, such as immunological measurements, should be reserved to research procedures related to the investigation of inflammatory network. However, due to the lack of standardization of parameters in the monitoring of NPT response, its reproducibility remains to be defined. The main problem includes the great variability of the responses in each patient and between patients. Although this is an important limitation, concerning NPT response interpretation, the symptom score has been used in the description of positive criteria to NPT response.

As pointed out above, many authors use the symptom score as a method of monitoring and criteria for positivity in response to the NPT. According to the great variability in each patient and between patients, it has been assumed the absence of pattern of response to the NPT. The attempt to standardize this methodology was characterized by the symptom score quantification, through the use of symptoms scaling. One of the most important limitations of this symptoms scaling, is the overemphasis on nasal obstruction, since firstly not all patients value the perception of this symptom, and secondly, when it is present, it can result from concomitant obstructive and inflammatory causes. Besides there is no clinical pattern of response to NPT, our data showed a response profile, which can not be accepted as standard, but it can be useful in monitoring the NPT, namely in the evaluation of the dynamics of the response to NPT, as described bellow.

#### **5.1 Clinical symptom score pattern**

In our experience, the symptom score has supported the positivity of NPT. We analysed that the most frequent and intense symptoms occurred within the first 30 minutes after NPT, agreeing to immediate phase of allergic reaction. From all the studies conducted by our group, we did not observe a clinical score symptom pattern. However, we describe a clinical symptom score profile, which was frequent and was characterized by the presence of nasal and extra-nasal symptoms within the first 30 minutes, with a peak at 5 minutes.

Fig. 2. Score symptoms after *Parietaria judaica* specific nasal provocation test (NPT) (legend: total score; nasal score; non-nasal score).

Indeed, in a group of patients allergic to *Parietaria judaica*, as described above, specific NPT was performed and a symptom score was recorded. The figure 2 presents the total, nasal and non-nasal symptom scores. The higher total score of symptoms was recorded at the fifth minute with progressively decreasing symptoms till 30 minutes and then till 6 hours. Each nasal symptom followed this pattern. The non-nasal symptoms showed a different pattern, having a lower score, with similar values at both the fifth and the 30th minutes, followed by a

rhinomanometry and acoustic rhinometry. The other methods, such as immunological measurements, should be reserved to research procedures related to the investigation of inflammatory network. However, due to the lack of standardization of parameters in the monitoring of NPT response, its reproducibility remains to be defined. The main problem includes the great variability of the responses in each patient and between patients. Although this is an important limitation, concerning NPT response interpretation, the

As pointed out above, many authors use the symptom score as a method of monitoring and criteria for positivity in response to the NPT. According to the great variability in each patient and between patients, it has been assumed the absence of pattern of response to the NPT. The attempt to standardize this methodology was characterized by the symptom score quantification, through the use of symptoms scaling. One of the most important limitations of this symptoms scaling, is the overemphasis on nasal obstruction, since firstly not all patients value the perception of this symptom, and secondly, when it is present, it can result from concomitant obstructive and inflammatory causes. Besides there is no clinical pattern of response to NPT, our data showed a response profile, which can not be accepted as standard, but it can be useful in monitoring the NPT, namely in the evaluation of the

In our experience, the symptom score has supported the positivity of NPT. We analysed that the most frequent and intense symptoms occurred within the first 30 minutes after NPT, agreeing to immediate phase of allergic reaction. From all the studies conducted by our group, we did not observe a clinical score symptom pattern. However, we describe a clinical symptom score profile, which was frequent and was characterized by the presence of nasal

**5 minutes 30 minutes 6 hours**

Fig. 2. Score symptoms after *Parietaria judaica* specific nasal provocation test (NPT) (legend:

Indeed, in a group of patients allergic to *Parietaria judaica*, as described above, specific NPT was performed and a symptom score was recorded. The figure 2 presents the total, nasal and non-nasal symptom scores. The higher total score of symptoms was recorded at the fifth minute with progressively decreasing symptoms till 30 minutes and then till 6 hours. Each nasal symptom followed this pattern. The non-nasal symptoms showed a different pattern, having a lower score, with similar values at both the fifth and the 30th minutes, followed by a

and extra-nasal symptoms within the first 30 minutes, with a peak at 5 minutes.

symptom score has been used in the description of positive criteria to NPT response.

dynamics of the response to NPT, as described bellow.

**5.1 Clinical symptom score pattern**

total score; nasal score; non-nasal score).

**before NPT**

decline till 6 hours. Looking at the score of each nasal symptom (Figure 3), except for nasal obstruction, all of them followed the response pattern of total symptoms score, with a peak of symptoms at the fifth minute. Sneezing was the predominant symptom at the fifth minute, while nasal obstruction was the predominant symptom at the 30th minute and the sixth hour.

Fig. 3. Nasal symptom score after specific *Parietaria judaica* nasal provocation test (NPT): evolution of each nasal symptom (Legend: nasal congestion; pruritus; sneezing; rhinorrea).

In another study mentioned above, the *Dermatophagoides pteronyssinus* specific NPT were performed in 34 children with OME. Those who had positive NPT, showed a response dynamics characterized by a rapid increase of symptoms score till a peak at the 5th minute (monitored till 1 hour), as shown in figure 4. Looking at the score of each nasal symptom (Figure 5), except for nasal pruritus, all followed the response pattern of total symptoms score, with the peak of symptoms at the fifth minute.

Fig. 4. Score symptoms after *Dermatophagoides pteronyssinus* specific nasal provocation test (NPT); (Legend: total score; nasal score; non-nasal score).

Nasal Provocation Test in the Diagnosis of Allergic Rhinitis 169

Fig. 6. Symptom scores reported during natural exposure and after nasal provocation test

**5.2 Comparison of** *Dermatophagoides pteronyssinus* **nasal provocation test** *versus*

describing the clinical pattern of NPT and CPT responses, neither about its comparison.

Our aim was to compare the dynamics of clinical responses induced by NPT and CPT, using

We studied two groups of voluntary adult patients, referred to our outpatient Immunoallergy Department, with *Dermatophagoides pteronyssinus* (*Dp*) allergic rhinitis/rhinoconjunctivitis with or without associated bronchial asthma, according to ARIA (Bousquet & ARIA Workshop Group, 2001) and GINA guidelines, respectively. All patients were clinically stable at the time of the study. Patients with past or ongoing immunotherapy for *Dermatophagoides*, an exacerbation of allergic disease or a respiratory tract infection in the last month, a nasal surgery in the last 3 months or nasal pathology such as polyps or a deviated nasal septum, were excluded. H1-antihistamines and costicosteroids, either nasal

Our group and others authors have been using clinical scores to evaluate NPT response. According to our findings described previously, in respect to the symptoms scores pattern in response to NPT, we conducted a study to characterize the clinical response to NPT comparing to conjunctiva provocation test (CPT). As CPT is easy to perform and systemic reactions are uncommon, some authors have studied the concordance between NPT and CPT in the diagnosis of allergic rhinitis (Andersen et al, 1996; Leonardi et al, 1993; Malmberg et al, 1978; Petersson et al, 1986; Riechelmann et al, 2003) and asthma (Mosbech et al, 1987) using clinical score symptoms and/or objective methods. However, we are not aware of any publication

**NPT**

Nasal congestion and pruritus were always reported and the latest had the higher recorded value (2.4±0.5)

The highest nasal recorded value during **NPT** was 6.4±2.19

Nasal congestion was always reported and it had the highest recorded value (2.8±0.35)

During **natural exposure**, the nasal total score was 6.2±2.05

**Natural exposure**

**conjunctiva provocation test**

a clinical score system.

**<IP_ADDRESS> Subjects**

**5.2.1 Material and methods**

(NPT).

Fig. 5. Nasal symptom score after specific *Dermatophagoides pteronyssinus* nasal provocation test (NPT): evolution of each nasal symptom (Legend: nasal congestion; pruritus; sneezing; rhinorrea).

Beyond the description of symptoms score obtained during NPT, it is also important to compare them with the usual symptoms described by the patient. This looks particularly relevant in the diagnosis of local allergic rhinitis.

In our study related to local allergic rhinitis diagnosis, we included 15 patients with typical clinical symptoms of perennial rhinitis, negative skin prick test to common aeroallergens and negative specific IgE, as mentioned above (Loureiro G et al, 2011). The patients had an average age of 22.214.8 years, 77.7% were female. A *Dermatophagoides pteronyssinus* specific NPT was performed with clinical monitoring. Total nasal symptom scores were assessed using a validated questionnaire and a positive challenge was considered if a score of five or greater was recorded. NPT supported the diagnosis of local allergic rhinitis in a group of patients previously diagnosed with "non-allergic rhinitis". They presented a period of symptoms evolution of 5.373.9 years. The symptom scores reported during natural exposure and after NPT are shown in figure 6. During natural exposure, the nasal total score was 6.22.05. Nasal congestion was always reported and it had the highest recorded value (2.80.35). The highest nasal recorded value during NPT was 6.42.19. Nasal congestion and pruritus were always reported and this second symptom had the higher recorded value (2.40.5). None of the 15 patients had conjunctivitis or asthma. Furthermore, in the 8 patients that had positive NPT, extra-nasal symptoms were recorded, namely conjunctival symptoms, oropharyngeal pruritus, cough and dyspnea, although with lower values.

Concerning the occurrence of non-nasal symptoms, the major non-nasal symptoms observed were those localized in the conjunctiva, followed by oropharyngeal pruritus. Dyspnea and cough were recognized rarely. Non-nasal symptoms were documented in 20 up to 100% of the positive NPT performed, considering the different studies conducted in our Immunoallergy Department.

**5th minute**

Fig. 5. Nasal symptom score after specific *Dermatophagoides pteronyssinus* nasal provocation test (NPT): evolution of each nasal symptom (Legend: nasal congestion; pruritus;

Beyond the description of symptoms score obtained during NPT, it is also important to compare them with the usual symptoms described by the patient. This looks particularly

In our study related to local allergic rhinitis diagnosis, we included 15 patients with typical clinical symptoms of perennial rhinitis, negative skin prick test to common aeroallergens and negative specific IgE, as mentioned above (Loureiro G et al, 2011). The patients had an average age of 22.214.8 years, 77.7% were female. A *Dermatophagoides pteronyssinus* specific NPT was performed with clinical monitoring. Total nasal symptom scores were assessed using a validated questionnaire and a positive challenge was considered if a score of five or greater was recorded. NPT supported the diagnosis of local allergic rhinitis in a group of patients previously diagnosed with "non-allergic rhinitis". They presented a period of symptoms evolution of 5.373.9 years. The symptom scores reported during natural exposure and after NPT are shown in figure 6. During natural exposure, the nasal total score was 6.22.05. Nasal congestion was always reported and it had the highest recorded value (2.80.35). The highest nasal recorded value during NPT was 6.42.19. Nasal congestion and pruritus were always reported and this second symptom had the higher recorded value (2.40.5). None of the 15 patients had conjunctivitis or asthma. Furthermore, in the 8 patients that had positive NPT, extra-nasal symptoms were recorded, namely conjunctival

symptoms, oropharyngeal pruritus, cough and dyspnea, although with lower values.

Concerning the occurrence of non-nasal symptoms, the major non-nasal symptoms observed were those localized in the conjunctiva, followed by oropharyngeal pruritus. Dyspnea and cough were recognized rarely. Non-nasal symptoms were documented in 20 up to 100% of the positive NPT performed, considering the different studies conducted in

**30th minute** **1 hour**

**0**

our Immunoallergy Department.

**before NPT**

relevant in the diagnosis of local allergic rhinitis.

**1st minute**

**1**

**2**

**3**

**0.5**

sneezing; rhinorrea).

**1.5**

**2.5**

Fig. 6. Symptom scores reported during natural exposure and after nasal provocation test (NPT).

#### **5.2 Comparison of** *Dermatophagoides pteronyssinus* **nasal provocation test** *versus* **conjunctiva provocation test**

Our group and others authors have been using clinical scores to evaluate NPT response. According to our findings described previously, in respect to the symptoms scores pattern in response to NPT, we conducted a study to characterize the clinical response to NPT comparing to conjunctiva provocation test (CPT). As CPT is easy to perform and systemic reactions are uncommon, some authors have studied the concordance between NPT and CPT in the diagnosis of allergic rhinitis (Andersen et al, 1996; Leonardi et al, 1993; Malmberg et al, 1978; Petersson et al, 1986; Riechelmann et al, 2003) and asthma (Mosbech et al, 1987) using clinical score symptoms and/or objective methods. However, we are not aware of any publication describing the clinical pattern of NPT and CPT responses, neither about its comparison. Our aim was to compare the dynamics of clinical responses induced by NPT and CPT, using

#### **5.2.1 Material and methods <IP_ADDRESS> Subjects**

a clinical score system.

We studied two groups of voluntary adult patients, referred to our outpatient Immunoallergy Department, with *Dermatophagoides pteronyssinus* (*Dp*) allergic rhinitis/rhinoconjunctivitis with or without associated bronchial asthma, according to ARIA (Bousquet & ARIA Workshop Group, 2001) and GINA guidelines, respectively. All patients were clinically stable at the time of the study. Patients with past or ongoing immunotherapy for *Dermatophagoides*, an exacerbation of allergic disease or a respiratory tract infection in the last month, a nasal surgery in the last 3 months or nasal pathology such as polyps or a deviated nasal septum, were excluded. H1-antihistamines and costicosteroids, either nasal

Nasal Provocation Test in the Diagnosis of Allergic Rhinitis 171

Demographical and clinical data are presented in Table 3. Table 4 shows the number of patients that presented nasal and ocular responses at the 5th minute, induced by NPT and CPT, as well as the number of positive challenges at the 1st and the 5th minutes. A progressive increase in clinical score was observed in both provocations. The NPT progressive response was linear while for the CPT it was exponential, as shown in figures 7. CPT response was stronger than NPT at the 5th minute, achieving borderline significance

(a) (b) Fig. 7. Dynamics of symptoms score in response to: A – NPT (Linear progression); B – CPT (Exponential progression) Legend: - Total Symptom score; - Nasal symptom score; -

The most frequent symptoms were nasal obstruction, itchy ear/throat and itchy nose, for NPT, and ocular hyperaemia and burning eyes, for CPT in all patients. In NPT, nasal obstruction was observed in 100% of the group. CPT induced ocular hyperaemia and burning eyes in all patients. There were neither bronchial symptoms nor systemic reactions

The highest scores were reached by nasal obstruction and rhinorrhea in NPT and by ocular hyperaemia in CPT. The average intensity of each sign/symptom at the 5th minute is shown

n 21 21 Average age (years) 28.0 9.0 28.1 5.7 Gender ♀ (%) 57.1 66.7

Associated asthma (%) 42.8 90.5

Disease evolution (years) 13 10 12.3 8.5 Table 3. Demographical and clinical data of patients submitted to NPT and CPT (Legend:

NPT – nasal provocation test; CPT – conjunctiva provocation test)

**before CPT 1st minute 5th minute**

**NPT CPT**

16 5

8.6 3.6 36.3 37.2

20 1

6.5 2.1 29 24.9

(p=0.05). Clinical score results for NPT and CPT are shown in Table 5.

**1st minute 5th minute 0**

**5.2.2 Results**

**-1**

in figure 8.

Rhinitis (n)

Rhinoconjunctivitis (n)

Cutaneous reactivity to Dp (mm) Specific IgE to Dp (KU/L)

**before NPT**

Non-nasal symptom score)

in any of the provocation tests.

**4**

**9**

**14**

or oral, were withheld for 2 weeks and 4 weeks prior to the challenge test, respectively. All patients underwent the challenge between January and February of 2009, a period of low natural exposure to mites in Portugal. A *Dp* NPT was performed in 21 patients and the conjunctival provocation test (CPT) was performed in the other 21 patients. The local ethics committee approved the study and all the participants gave written informed consent before entry. A respiratory function test (pletismography using *Master screen Body Jaeger*®) was performed by all the participants, before specific provocation tests, with all presenting a baseline FEV1 80% and FEV1/FVC 80. After provocation, all patients were asked for the presence of dyspnoea, thoracic oppression, wheezing or cough.

#### **<IP_ADDRESS> Specific nasal and conjuctival provocation tests**

A skin prick test aqueous extract of *Dp* with a 5 mg/ml concentration (23 g/ml of *Der p* 1, BialAristegui, Bilbao, Spain), with 1/1, 1/10, 1/100 and 1/1000 dilutions were performed; negative and positive controls were performed in all patients, according to standardized procedures (Dreborg & Frew A, 1993). The concentration used to specific provocation was the minimum that induced a prick test wheal at least equal to that induced by histamine, which curiously was the 1/10 dilution in all patients. Specific NPT with *Dp* extract were performed in the morning and after an adaptation to room temperature for 30 minutes, in both groups. NPT was performed with unilateral nasal application of 2 consecutives puffs (total volume of 160 l) of the *Dp* extract to the inferior nasal turbinate of the less congested nostril, using a nasal applicator spraying and patients were asked to perform apnoea during the allergen spraying. CPT consisted in unilateral ocular application of 1 drop (50 l) of the *Dp* extract in the inferior and external quadrant of the bulbar conjunctiva. Nasal and eye symptoms were recorded at the 1st and 5th minutes after specific provocation tests, using a clinical score system to assess the response (Linder A, 1988).

#### **<IP_ADDRESS> Clinical score scaling**

Clinical responses were evaluated using a nasal clinical score (NCS) and an ocular clinical score (OCS), at the 1st and the 5th minutes. An adaptation of the previously used NCS (Linder A, 1988) and OCS (Mortemousque, 2007) were applied. A total clinical score (TCS), representing the sum of NCS (range: 0-15) and OCS (range: 0-13) was also used, ranging from 0 to 28 points. Rhinorrhea, sneezing, itchy nose, itchy ear/throat, nasal obstruction, watery eyes, redness of eyes and burning of eyes were rated on a scale from 0 to 3 points (0, none; 1, mild; 2, moderate; 3, severe). Itchy eyes were scored from 0 to 4 points (0, none; 1, mild; 2, moderate; 3, severe; 4, very severe). A positive response to NPT was considered when NCS 3 (Linder A, 1988) and to CPT when OCS 5 (Mortemousque, 2007). Clinical evaluation was interrupted after the 5th minute to collect humours for further investigation to determine inflammatory markers within a research investigation of immunologic mechanisms in allergic disease (Pereira, 2011, *in press*).

#### **<IP_ADDRESS> Statistical analysis**

Statistical analysis were performed using SPSS® Statistics 17.0 software. Comparisons between NPT and CPT were studied using Chi-Square test. Intra-groups differences between the 1st and the 5th minutes after provocation were analyzed using a Mann-Whitney U-test. A statistical significant difference was assumed with p < 0.05.

#### **5.2.2 Results**

170 Allergic Rhinitis

or oral, were withheld for 2 weeks and 4 weeks prior to the challenge test, respectively. All patients underwent the challenge between January and February of 2009, a period of low natural exposure to mites in Portugal. A *Dp* NPT was performed in 21 patients and the conjunctival provocation test (CPT) was performed in the other 21 patients. The local ethics committee approved the study and all the participants gave written informed consent before entry. A respiratory function test (pletismography using *Master screen Body Jaeger*®) was performed by all the participants, before specific provocation tests, with all presenting a baseline FEV1 80% and FEV1/FVC 80. After provocation, all patients were asked for the

A skin prick test aqueous extract of *Dp* with a 5 mg/ml concentration (23 g/ml of *Der p* 1, BialAristegui, Bilbao, Spain), with 1/1, 1/10, 1/100 and 1/1000 dilutions were performed; negative and positive controls were performed in all patients, according to standardized procedures (Dreborg & Frew A, 1993). The concentration used to specific provocation was the minimum that induced a prick test wheal at least equal to that induced by histamine, which curiously was the 1/10 dilution in all patients. Specific NPT with *Dp* extract were performed in the morning and after an adaptation to room temperature for 30 minutes, in both groups. NPT was performed with unilateral nasal application of 2 consecutives puffs (total volume of 160 l) of the *Dp* extract to the inferior nasal turbinate of the less congested nostril, using a nasal applicator spraying and patients were asked to perform apnoea during the allergen spraying. CPT consisted in unilateral ocular application of 1 drop (50 l) of the *Dp* extract in the inferior and external quadrant of the bulbar conjunctiva. Nasal and eye symptoms were recorded at the 1st and 5th minutes after specific provocation tests, using a clinical score system to assess the

Clinical responses were evaluated using a nasal clinical score (NCS) and an ocular clinical score (OCS), at the 1st and the 5th minutes. An adaptation of the previously used NCS (Linder A, 1988) and OCS (Mortemousque, 2007) were applied. A total clinical score (TCS), representing the sum of NCS (range: 0-15) and OCS (range: 0-13) was also used, ranging from 0 to 28 points. Rhinorrhea, sneezing, itchy nose, itchy ear/throat, nasal obstruction, watery eyes, redness of eyes and burning of eyes were rated on a scale from 0 to 3 points (0, none; 1, mild; 2, moderate; 3, severe). Itchy eyes were scored from 0 to 4 points (0, none; 1, mild; 2, moderate; 3, severe; 4, very severe). A positive response to NPT was considered when NCS 3 (Linder A, 1988) and to CPT when OCS 5 (Mortemousque, 2007). Clinical evaluation was interrupted after the 5th minute to collect humours for further investigation to determine inflammatory markers within a research investigation of immunologic

Statistical analysis were performed using SPSS® Statistics 17.0 software. Comparisons between NPT and CPT were studied using Chi-Square test. Intra-groups differences between the 1st and the 5th minutes after provocation were analyzed using a Mann-Whitney

presence of dyspnoea, thoracic oppression, wheezing or cough.

**<IP_ADDRESS> Specific nasal and conjuctival provocation tests**

mechanisms in allergic disease (Pereira, 2011, *in press*).

U-test. A statistical significant difference was assumed with p < 0.05.

response (Linder A, 1988). **<IP_ADDRESS> Clinical score scaling**

**<IP_ADDRESS> Statistical analysis**

Demographical and clinical data are presented in Table 3. Table 4 shows the number of patients that presented nasal and ocular responses at the 5th minute, induced by NPT and CPT, as well as the number of positive challenges at the 1st and the 5th minutes. A progressive increase in clinical score was observed in both provocations. The NPT progressive response was linear while for the CPT it was exponential, as shown in figures 7. CPT response was stronger than NPT at the 5th minute, achieving borderline significance (p=0.05). Clinical score results for NPT and CPT are shown in Table 5.

Fig. 7. Dynamics of symptoms score in response to: A – NPT (Linear progression); B – CPT (Exponential progression) Legend: - Total Symptom score; - Nasal symptom score; - Non-nasal symptom score)

The most frequent symptoms were nasal obstruction, itchy ear/throat and itchy nose, for NPT, and ocular hyperaemia and burning eyes, for CPT in all patients. In NPT, nasal obstruction was observed in 100% of the group. CPT induced ocular hyperaemia and burning eyes in all patients. There were neither bronchial symptoms nor systemic reactions in any of the provocation tests.

The highest scores were reached by nasal obstruction and rhinorrhea in NPT and by ocular hyperaemia in CPT. The average intensity of each sign/symptom at the 5th minute is shown in figure 8.

Table 3. Demographical and clinical data of patients submitted to NPT and CPT (Legend: NPT – nasal provocation test; CPT – conjunctiva provocation test)

Total 1st min 5th min 5th - 1st

Nasal (NCS) 1st min 5th min

Ocular (OCS) 1st min 5th min

– conjunctiva provocation test; ns – not significant).

Nasal Provocation Test in the Diagnosis of Allergic Rhinitis 173

5.2 3.8 9.9 4.4 4.6 4.57

4.28 2.6 8.29 2.9

0.95 1.8 1.57 2.3

Table 5. Clinical score results for NPT and CPT (Legend: NPT – nasal provocation test; CPT

all symptoms, are easy and costless to apply in clinical practice. The validity and reproducibility of CPT based on clinical score systems were demonstrated in several studies

According to our findings, we can describe a dynamic response profile to specific provocation. In our study, NPT response at the 1st minute was faster than CPT (*p*=0.005), with 15/21 patients presenting a positive NPT *versus* 6/21 patients with positive CPT. We speculate that this can eventually be explained by the existence of particular characteristics in nasal and ocular mucosa, resulting in differences related to the contact with the allergen and/or the time response of type I hypersensitivity. The NPT progressive response was

On the other hand, CPT response was stronger at the 5th minute when comparing to NPT, achieving borderline significance (*p*=0.05). This corroborates other results related to the evaluation of patient discomfort of NPT *versus* CPT using a visual-analogue scale, with a higher discomfort being appointed to CPT (Riechelmann et al, 2003). Apparently, these results are different from the study of Malmberg et al, 1978, in which the conjunctiva of 55% of the patients that underwent both NPT and CPT, using sequentially diluted allergen solutions, was less sensitive to allergen challenge than nasal mucosa. However, the intensity of the positive CPT response was not described in this study. Our patients submitted to CPT had higher specific IgE values, but it is unlikely that this could explain the higher intensity symptoms score. The absence of a direct correlation between the degree of allergen

As expected by direct allergen exposure, the higher intensity of nasal response was induced

At the 5th minute, procedures to collect secretions were performed, and consequently the clinical evaluation of the response to specific provocation tests was disrupted. However, patients were clinically monitored till 4th hour. Interestingly, after the 5th minute, the intensity of the conjunctival response rapidly decreased while a similar intensity of nasal response persisted for a longer period. This data is not shown because the procedures for

by NPT, while CPT was responsible for the higher intensity of ocular response.

(Abelson et al, 1990; Moller et al, 1984; Mortemousque, 2007; Rimas et al, 1992).

linear whereas CPT one was exponential, till the 5th minute of response.

sensitization and the severity of clinical symptoms is well known.

collection of secretions could alter the dynamic of response.

**NPT CPT** *p*

4.7 3.6 12.7 4.4 8.0 3.87

1.24 2.1 4.95 2.8

> 3.4 3 7.7 3

ns ns (0.05) 0.011

<0.0001 0.001

<0.0001 <0.0001

Table 4. Frequency of nasal and ocular symptoms at the 5th minute and NPT and CPT outcomes at the 1st and the 5th minutes (Legend: NPT – nasal provocation test; CPT – conjunctiva provocation test; ns - not significant).

Comparing NPT and CPT, in the first one the response was faster at the 1st minute (p=0.005) while for CPT it was stronger at the 5th minute (p=0.05).

Although the inoculation of allergen was unilateral, NPT induced bilateral nasal symptoms in 100% and bilateral ocular symptoms in 47.6%. On the other hand, CPT induced unilateral ocular symptoms in 100% and bilateral nasal symptoms in 95.2%. There were neither bronchial symptoms nor systemic reactions.

Fig. 8. The average intensity of each sign/symptom at the 5th minute; (Legend: - Nasal provocation test; - Conjunctiva provocation test)

#### **5.2.3 Discussion**

Although the importance of the objective monitoring of specific provocation tests is unquestionable, its applicability in clinical practice is not always possible. Usually it is limited to the evaluation of only one symptom, such as nasal patency by nasal peak flow, acoustic rhinometry and/or rhinomanometry (Nathan et al, 2005); however it is not always the most perceived symptom by patients. Clinical scoring systems, even though more subjective, reflect

Table 4. Frequency of nasal and ocular symptoms at the 5th minute and NPT and CPT outcomes at the 1st and the 5th minutes (Legend: NPT – nasal provocation test; CPT –

Comparing NPT and CPT, in the first one the response was faster at the 1st minute (p=0.005)

Although the inoculation of allergen was unilateral, NPT induced bilateral nasal symptoms in 100% and bilateral ocular symptoms in 47.6%. On the other hand, CPT induced unilateral ocular symptoms in 100% and bilateral nasal symptoms in 95.2%. There were neither

21 (100%) 10 (47.6%)

> 15 21

0123

Fig. 8. The average intensity of each sign/symptom at the 5th minute; (Legend: - Nasal

Although the importance of the objective monitoring of specific provocation tests is unquestionable, its applicability in clinical practice is not always possible. Usually it is limited to the evaluation of only one symptom, such as nasal patency by nasal peak flow, acoustic rhinometry and/or rhinomanometry (Nathan et al, 2005); however it is not always the most perceived symptom by patients. Clinical scoring systems, even though more subjective, reflect

Nasal response at 5th min Ocular response at 5th min

1st min 5th min

Number of positive challenges:

conjunctiva provocation test; ns - not significant).

bronchial symptoms nor systemic reactions.

provocation test; - Conjunctiva provocation test)

ocular itching

**5.2.3 Discussion**

lacrimation

chemosis

ocular erythema

palate itching

nasal itching

rhinorrea

nasal obstruction

sneeze

while for CPT it was stronger at the 5th minute (p=0.05).

**NPT CPT** *p*

20 (95.2%) 21 (100%)

6

<sup>21</sup>0.005

ns 0.0001

Table 5. Clinical score results for NPT and CPT (Legend: NPT – nasal provocation test; CPT – conjunctiva provocation test; ns – not significant).

all symptoms, are easy and costless to apply in clinical practice. The validity and reproducibility of CPT based on clinical score systems were demonstrated in several studies (Abelson et al, 1990; Moller et al, 1984; Mortemousque, 2007; Rimas et al, 1992).

According to our findings, we can describe a dynamic response profile to specific provocation. In our study, NPT response at the 1st minute was faster than CPT (*p*=0.005), with 15/21 patients presenting a positive NPT *versus* 6/21 patients with positive CPT. We speculate that this can eventually be explained by the existence of particular characteristics in nasal and ocular mucosa, resulting in differences related to the contact with the allergen and/or the time response of type I hypersensitivity. The NPT progressive response was linear whereas CPT one was exponential, till the 5th minute of response.

On the other hand, CPT response was stronger at the 5th minute when comparing to NPT, achieving borderline significance (*p*=0.05). This corroborates other results related to the evaluation of patient discomfort of NPT *versus* CPT using a visual-analogue scale, with a higher discomfort being appointed to CPT (Riechelmann et al, 2003). Apparently, these results are different from the study of Malmberg et al, 1978, in which the conjunctiva of 55% of the patients that underwent both NPT and CPT, using sequentially diluted allergen solutions, was less sensitive to allergen challenge than nasal mucosa. However, the intensity of the positive CPT response was not described in this study. Our patients submitted to CPT had higher specific IgE values, but it is unlikely that this could explain the higher intensity symptoms score. The absence of a direct correlation between the degree of allergen sensitization and the severity of clinical symptoms is well known.

As expected by direct allergen exposure, the higher intensity of nasal response was induced by NPT, while CPT was responsible for the higher intensity of ocular response.

At the 5th minute, procedures to collect secretions were performed, and consequently the clinical evaluation of the response to specific provocation tests was disrupted. However, patients were clinically monitored till 4th hour. Interestingly, after the 5th minute, the intensity of the conjunctival response rapidly decreased while a similar intensity of nasal response persisted for a longer period. This data is not shown because the procedures for collection of secretions could alter the dynamic of response.

Nasal Provocation Test in the Diagnosis of Allergic Rhinitis 175

We would like to acknowledge Dr Borja Bartolomé, Bial Aristegui, I&D Department, Bilbao, Spain; Dr António Martinho & Dr Artur Paiva, PhD, Histocompatibility Center, Coimbra,

Abelson, MB.; Chambers, WA. & Smith, LM. (1990). Conjunctival allergen challenge. A

Anderson, DF.; McGill, JI. & Roche, WR. (1996). Improving the safety of conjunctival

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Baroody, FM.; Foster, KA.; Markaryan, A.; deTineo, M. & Naclerio, RM. (2008). Nasal Ocular

Baroody, FM. (2011). How nasal function influences the eyes, ears, sinuses and lungs. *Proc Am Thorac Soc*, Vol.8, No.1, (March 2011), pp. 53-61, ISSN 1546-3222 Bousquet, J.; Van Cauwenberge, P.; Khaltaev, N.; Aria Workshop Group & World Health

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**7. Acknowledgements**

Portugal.

**8. References**

Even though the allergen was unilaterally inoculated, NPT induced bilateral nasal symptoms in 100% and bilateral ocular symptoms in 47.6%. On the other hand, CPT induced unilateral ocular symptoms in 100% and bilateral nasal symptoms in 95.2%. This is in accordance with previous studies and can be explained by different mechanisms mentioned above (Section 4. NPT as a diagnostic approach in respiratory allergy). An additional explanation for the higher number of patients with nasal symptoms induced by CPT, when comparing with the number of patients in whom NPT induced ocular symptoms, is the direct contact of the inoculated allergen with the nasal mucosa, through its passage via naso-lacrimal duct.

This study describes, for the first time to our knowledge, the clinical patterns of NPT and CPT responses, using a clinical score system. NPT is faster than CPT and has a linear progression, while CPT has an exponential progression and has a stronger response. The induction of both nasal and ocular responses by NPT or CPT, corroborates the systemically response triggered by local allergen application. Although both methodologies can elicit extra-local symptoms, these are safe procedures. Finally, these data support the applicability of CPT in the diagnosis of allergic rhinitis, even in the absence of ocular signs/symptoms, surpassing some NPT limitations (such as nasal polyps or deviated nasal septum) and decreasing specific challenge risk.

#### **6. Conclusion**

The specific provocation tests have been widely used in the investigation of pathophysiological mechanisms, immunological and therapeutic aspects of allergic disease, since they mimic the response to allergen exposure, under controlled conditions. It is well known that NPT has limitations, but it has been helpful to a better clarification of the underlying mechanisms of allergic reaction, and also to recognize the systemic framework of allergic disease. The usefulness of NPT is focused in the diagnosis of allergic rhinitis itself, but it has also a relevant role in the diagnosis of allergic respiratory disease. The upper and lower airways do not exist as anatomically and functionally distinct areas. There are important relationships between both the nose and the paranasal sinuses, and asthma. These epidemiological, clinical and immunopathologic concordance between allergic rhinitis and asthma supports the concept of bidirectional "unified airway" respiratory inflammation model. Multiple evidence supports a close interaction and influence of the nose on contiguous and distant organs via neural reflex and systemic inflammatory processes.

In clinical practice, NPT plays a central role in the diagnosis of allergic rhinitis in some circumstances, as described. This is the only method that could establish the correct aetiology of the allergic disease, namely local allergic rhinitis and occupational rhinitis. The specific therapeutic implications emphasize the attempt to reach the most complete diagnostic approach.

The monitoring of the response to NPT is not standardized, but several parameters have been used, for example symptom scores. Our data suggest that the clinical symptom pattern to NPT develops has a dynamic response which is characterized by a linear progression of symptoms intensity till a 5th minute peak. The prevalence of non-nasal symptoms had a great variability in the studies performed by our group. Those symptoms had a lower score comparing to nasal symptoms. In our opinion, the symptom score is a valuable method to monitor the NPT response.

#### **7. Acknowledgements**

We would like to acknowledge Dr Borja Bartolomé, Bial Aristegui, I&D Department, Bilbao, Spain; Dr António Martinho & Dr Artur Paiva, PhD, Histocompatibility Center, Coimbra, Portugal.

#### **8. References**

174 Allergic Rhinitis

Even though the allergen was unilaterally inoculated, NPT induced bilateral nasal symptoms in 100% and bilateral ocular symptoms in 47.6%. On the other hand, CPT induced unilateral ocular symptoms in 100% and bilateral nasal symptoms in 95.2%. This is in accordance with previous studies and can be explained by different mechanisms mentioned above (Section 4. NPT as a diagnostic approach in respiratory allergy). An additional explanation for the higher number of patients with nasal symptoms induced by CPT, when comparing with the number of patients in whom NPT induced ocular symptoms, is the direct contact of the inoculated allergen with the nasal mucosa, through its

This study describes, for the first time to our knowledge, the clinical patterns of NPT and CPT responses, using a clinical score system. NPT is faster than CPT and has a linear progression, while CPT has an exponential progression and has a stronger response. The induction of both nasal and ocular responses by NPT or CPT, corroborates the systemically response triggered by local allergen application. Although both methodologies can elicit extra-local symptoms, these are safe procedures. Finally, these data support the applicability of CPT in the diagnosis of allergic rhinitis, even in the absence of ocular signs/symptoms, surpassing some NPT limitations (such as nasal polyps or deviated nasal septum) and

The specific provocation tests have been widely used in the investigation of pathophysiological mechanisms, immunological and therapeutic aspects of allergic disease, since they mimic the response to allergen exposure, under controlled conditions. It is well known that NPT has limitations, but it has been helpful to a better clarification of the underlying mechanisms of allergic reaction, and also to recognize the systemic framework of allergic disease. The usefulness of NPT is focused in the diagnosis of allergic rhinitis itself, but it has also a relevant role in the diagnosis of allergic respiratory disease. The upper and lower airways do not exist as anatomically and functionally distinct areas. There are important relationships between both the nose and the paranasal sinuses, and asthma. These epidemiological, clinical and immunopathologic concordance between allergic rhinitis and asthma supports the concept of bidirectional "unified airway" respiratory inflammation model. Multiple evidence supports a close interaction and influence of the nose on contiguous and distant organs via neural reflex and systemic inflammatory processes. In clinical practice, NPT plays a central role in the diagnosis of allergic rhinitis in some circumstances, as described. This is the only method that could establish the correct aetiology of the allergic disease, namely local allergic rhinitis and occupational rhinitis. The specific therapeutic implications emphasize the attempt to reach the most complete

The monitoring of the response to NPT is not standardized, but several parameters have been used, for example symptom scores. Our data suggest that the clinical symptom pattern to NPT develops has a dynamic response which is characterized by a linear progression of symptoms intensity till a 5th minute peak. The prevalence of non-nasal symptoms had a great variability in the studies performed by our group. Those symptoms had a lower score comparing to nasal symptoms. In our opinion, the symptom score is a valuable method to

passage via naso-lacrimal duct.

decreasing specific challenge risk.

**6. Conclusion**

diagnostic approach.

monitor the NPT response.

Nasal Provocation Test in the Diagnosis of Allergic Rhinitis 177

Ganslmayer, M.; Spertini, F.; Rahm, F.; Terrien, MH.; Mosimann, B. & Leimgruber, A. (1999).

Gosepath, J.; Amedee, RG. & Mann, WJ. (2005). Nasal provocation testing as an international

Gotlib, T.; Samoliński, B. & Grzanka, A. (2005). Bilateral nasal allergen provocation

Gregory, LG.; Causton, B.; Murdoch, JR.; Mathie, SA.; O'Donnell, V.; Thomas, CP.; Priest,

Hammad, H.; Chieppa, M.; Perros, F.; Willart, MA.; Germain, RN. & Lambrecht, BN. (2009).

Hens, G.; Bobic, S.; Reekmans, K.; Ceuppens, JL. & Hellings, PW. (2007). Rapid systemic

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**11**

*Taiwan*

**Phototherapy for the Treatment**

*1Department of Otorhinolaryngology, Keelung Hospital*

*2School of Traditional Chinese Medicine, Chang Gung University*

*3Department of Biomedical Engineering, Chung-Yuan Christian University*

Allergic rhinitis (AR) is one of the most common allergic diseases, affecting 20% of the adult population and up to 40% of children (Salib et al., 2003). It is associated with decreased learning, performance and productivity at work and school, as well as a reduced quality of life. The detrimental effects of AR on quality of life (QOL) include fatigue, irritability, memory deficits, excessive daytime somnolence, and depression. The annual economic impact of AR is calculated to be between \$ 6.3 billion and \$ 7.9 billion without counting its detrimental effects on QOL (Fineman, 2002). Current therapeutic options such as allergen avoidance, medication and immunotherapy are far from ideal. It is important to develop an effective modality to relieve the symptom except for targeting the complexity of underlying

Phototherapy is the application of light to a pathological area to promote tissue regeneration, reduce inflammation and relieve pain. Several types of phototherapeutic devices are currently used for medical treatment using selected wavelengths and controlled dosage of irradiation. Significant suppression on the clinical symptoms of AR by the phototherapy treatment of ultraviolet (UV) and visible light was reported (Csoma, et al., 2004, 2006; Koreck, et al., 2005, 2007). Narrow-band red light phototherapy was found to markedly alleviate the clinical symptoms of AR (Neuman & Finkelstein, 1997). In addition to UV and visible light therapy, far infrared ray (FIR) therapy is also reported to have beneficial effects to patients with AR (Hu & Li, 2007). Photochemical effect is elicited using UV and visible light irradiation, whereas thermal effect is induced with FIR irradiation. Although different mechanisms are involved when light sources with different ranges of wavelengths are employed, phototherapy

This chapter is organized as follows. First, pathophysiology and traditional management of AR are briefly reviewed. Second, photobiology and phototherapy related to AR are summarized. Finally, the clinical outcomes of FIR therapy as well as red light acupoint

AR is defined as an abnormal inflammation of the membrane lining the nose, which is mediated by immunoglobulin E (IgE). The clinical symptoms of AR include sneezing,

represents a noninvasive, alternative intervention for the treatment of AR.

stimulation on patients with AR are described.

**2. Pathophysiology of allergic rhinitis**

**1. Introduction**

inflammatory mechanism of AR.

 **of Allergic Rhinitis**

Ko-Hsin Hu1,2,3 and Wen-Tyng Li3

## **Phototherapy for the Treatment of Allergic Rhinitis**

Ko-Hsin Hu1,2,3 and Wen-Tyng Li3 *1Department of Otorhinolaryngology, Keelung Hospital 2School of Traditional Chinese Medicine, Chang Gung University 3Department of Biomedical Engineering, Chung-Yuan Christian University Taiwan*

#### **1. Introduction**

182 Allergic Rhinitis

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allergen Der p 1 elevates the release of inflammatory cytokines and expression of adhesion molecules in co-culture of human eosinophils and bronchial epithelial cells. *Int Immunol*, Vol. 18, No. 8 (August 2006), pp. 1327–1335, ISSN 0953-8178 Yeo, SG.; Park, DC.; Eun, YG.; & Cha, C. (2007). The role of allergic rhinitis in the

development of otitis media with effusion: effect on Eustachian tube function. *Am J*

sensitizations to enzymes after occupational exposure: evaluation by skin prick test, RAST, and immunoblot. *Allergy,* Vol. 52, No. 9 (September 1997), pp. 928-934, ISSN Allergic rhinitis (AR) is one of the most common allergic diseases, affecting 20% of the adult population and up to 40% of children (Salib et al., 2003). It is associated with decreased learning, performance and productivity at work and school, as well as a reduced quality of life. The detrimental effects of AR on quality of life (QOL) include fatigue, irritability, memory deficits, excessive daytime somnolence, and depression. The annual economic impact of AR is calculated to be between \$ 6.3 billion and \$ 7.9 billion without counting its detrimental effects on QOL (Fineman, 2002). Current therapeutic options such as allergen avoidance, medication and immunotherapy are far from ideal. It is important to develop an effective modality to relieve the symptom except for targeting the complexity of underlying inflammatory mechanism of AR.

Phototherapy is the application of light to a pathological area to promote tissue regeneration, reduce inflammation and relieve pain. Several types of phototherapeutic devices are currently used for medical treatment using selected wavelengths and controlled dosage of irradiation. Significant suppression on the clinical symptoms of AR by the phototherapy treatment of ultraviolet (UV) and visible light was reported (Csoma, et al., 2004, 2006; Koreck, et al., 2005, 2007). Narrow-band red light phototherapy was found to markedly alleviate the clinical symptoms of AR (Neuman & Finkelstein, 1997). In addition to UV and visible light therapy, far infrared ray (FIR) therapy is also reported to have beneficial effects to patients with AR (Hu & Li, 2007). Photochemical effect is elicited using UV and visible light irradiation, whereas thermal effect is induced with FIR irradiation. Although different mechanisms are involved when light sources with different ranges of wavelengths are employed, phototherapy represents a noninvasive, alternative intervention for the treatment of AR.

This chapter is organized as follows. First, pathophysiology and traditional management of AR are briefly reviewed. Second, photobiology and phototherapy related to AR are summarized. Finally, the clinical outcomes of FIR therapy as well as red light acupoint stimulation on patients with AR are described.

#### **2. Pathophysiology of allergic rhinitis**

AR is defined as an abnormal inflammation of the membrane lining the nose, which is mediated by immunoglobulin E (IgE). The clinical symptoms of AR include sneezing,

Phototherapy for the Treatment of Allergic Rhinitis 185

Neuropeptides induce vasodilation, thus causing congestion (Howarth, 1997). Cytokine secretion upregulates the expression of adhesion molecules on the vascular endothelial cells, thereby enhancing the activation and adhesion of inflammatory cells. An increase in interleukin-4 (IL-4), IL-5, and granulocyte-macrophage colony-stimulating factor (GM-CSF)

Effective allergen avoidance can lead to substantial relief of symptoms. However, patients are still not able to avoid their confirmed allergens such as mites or atmospheric pollens under many circumstances. Medication to manipulate the release of mediators is the next step in the management of AR. Table 2 summarizes the classes of pharmacological therapies for the treatment of AR. The two major classes of medication are oral H1 antihistamines and intranasal corticosteroids. According to the guidelines, oral antihistamines are the first-line therapy which relieve sneezing and rhinorrhea (Bousquet, et al., 2001; van Cauwenberge, et al., 2000; Dykewicz, et al., 1998). Topical administration can further minimize systemic adverse effects of antihistamines. H1 antihistamines are often combined with a decongestant to reach sufficient efficacy for nasal congestion (Quraishi, et al., 2004). The new generation of antihistamines acts as inverse agonists that stabilize the inactive conformation of the receptors and reverses constitutive activity of receptors (Oppenheimer & Casale, 2002). Intranasal corticosteroids are recommended as first-line treatment for moderate and severe AR, which are effective in relieving symptoms such as sneezing, rhinorrhea, itching and congestion (Weiner, et al., 1998). Corticosteroids target the inflammatory mechanisms; therefore the amount of oral corticosteroids for long-term treatment should be carefully adjusted to avoid adverse effects such as osteoporosis and growth inhibition in children

Decongestants can reduce nasal obstruction and congestion by their vasoconstrictive action on α-adrenergic receptors. The application of intranasal decongestants may cause rhinitis medicamentosa. The adverse effects of oral decongestants include elevated blood pressure, tremor, tachycardia, loss of appetite, sleep disturbance. The long term use of decongestant is not recommended. (Quraishi, et al., 2004). Anticholinergics act as muscarinic receptor blocker which inhibits mucus secretion and subsequent rhinorrhea. Through inhibiting degranulation and neosynthesis of inflammatory mediators, mast cell stabilizers are shown to be effective in reducing symptoms of early inflammatory phase and are useful for preventive purposes (Al Suleimani & Walker, 2007). Leukotrienes inhibitors significantly reduce nasal blockade by inhibiting leukotriene synthesis or serving as antagonists for its receptors. Immunotherapy has the potential to provide a permanent cure for the disease. The proposed mechanisms for immunotherapy include suppression of IgE elevation, decrease in neutrophil and eosionophil activity, reduction in mast cell number, inhibition of T-lymphocyte proliferation (Jayasekera, et al., 2007; Pipet, et al., 2009). However, the technique is burdensome which requiring a lengthy series of injection and it may not be applicable to all patients (Naclerio, et al., 2002). Although most of the drugs are effective in treating certain symptoms of AR, they all have limitations due to their adverse effects. Due to the complex mechanisms involved in the AR, the ideal treatment for this disease has yet

is associated with a mucosal eosinophilia (Quraishi, et al., 2004).

**3. Traditional management of allergic rhinitis**

(Wilson, et al., 1998).

to be discovered.

itching of the nose, rhinorrhea and nasal congestion. Additionally, airway lining hypersensitivity, a loss of the sense of small and an inability to taste may occur. It has become progressively clear that it is a common comorbid condition with asthma, allergic conjunctivitis, sinusitis, otitis media, nasal polyposis and respiratory infections (Berrettini, et al., 1999; Skoner, 2000). Nasal obstruction can often be seen with pale nasal mucosa, enlarged turbinates, clear nasal secretions, and pharyngeal cobble-stoning upon physical examination (Al Suleimani & Walker, 2007). The diagnosis of AR was based on definite symptoms of nasal itching, rhinorrhea, sneezing, nasal obstruction or mouth breathing, as well as positive reactions to blood tests to antigens, such as house dust mite, cockroach, molds, feathers, grass pollen, weed pollens, sage pollen, and local tree pollens, etc. Criteria for positive skin prick test responses were a wheel of 3 mm or greater diameter with erythema of at least 5 mm. Histamine control skin test was read at 10 minutes, allergen and negative control skin tests were read at 15 minutes. The score for each symptom is usually registered on a four-grade scale- absent, slight, moderate or severe (Linder, 1988) as shown in Table 1.

Table 1. Scoring of symptoms for AR

Various mediators are associated to the pathophysiology of AR. For example, histamine plays a pivotal role in early allergic responses and also acts as stimulatory signal for cytokine production, expression of cell adhesion molecules and HLA class II antigens. Most of the effects of histamine in allergic disease are mediated through H1 receptors (Akdis & Blaser, 2003). Cysteinyl leukotrienes (CysLTs) increase nasal airway resistance and obstruction, and contribute to rhinorrhea via increased vascular permeability and mucus secretion (Okuda, et al., 1988). Prostaglandins cause congestion and rhinorrhea.

itching of the nose, rhinorrhea and nasal congestion. Additionally, airway lining hypersensitivity, a loss of the sense of small and an inability to taste may occur. It has become progressively clear that it is a common comorbid condition with asthma, allergic conjunctivitis, sinusitis, otitis media, nasal polyposis and respiratory infections (Berrettini, et al., 1999; Skoner, 2000). Nasal obstruction can often be seen with pale nasal mucosa, enlarged turbinates, clear nasal secretions, and pharyngeal cobble-stoning upon physical examination (Al Suleimani & Walker, 2007). The diagnosis of AR was based on definite symptoms of nasal itching, rhinorrhea, sneezing, nasal obstruction or mouth breathing, as well as positive reactions to blood tests to antigens, such as house dust mite, cockroach, molds, feathers, grass pollen, weed pollens, sage pollen, and local tree pollens, etc. Criteria for positive skin prick test responses were a wheel of 3 mm or greater diameter with erythema of at least 5 mm. Histamine control skin test was read at 10 minutes, allergen and negative control skin tests were read at 15 minutes. The score for each symptom is usually registered on a four-grade scale- absent, slight, moderate or severe (Linder, 1988) as shown

> Scoring of rhinorrhea 0: no nasal blowing

Scoring of smell impairment 0: no smell impairment

day

Various mediators are associated to the pathophysiology of AR. For example, histamine plays a pivotal role in early allergic responses and also acts as stimulatory signal for cytokine production, expression of cell adhesion molecules and HLA class II antigens. Most of the effects of histamine in allergic disease are mediated through H1 receptors (Akdis & Blaser, 2003). Cysteinyl leukotrienes (CysLTs) increase nasal airway resistance and obstruction, and contribute to rhinorrhea via increased vascular permeability and mucus secretion (Okuda, et al., 1988). Prostaglandins cause congestion and rhinorrhea.

impairment 3: anosmia

Scoring of sneezing 0: no sneezing

1:nasal blowing less than 5 episodes a day 2: nasal blowing 6-10 episodes a day

1: hyposmia with mild smell impairment 2: hyposmia with moderate smell

1: sneezing less than 5 episodes a day 2: sneezing 6-10 episodes a day

3: sneezing more than 10 episodes a day

3: nasal blowing more than 10 episodes a

in Table 1.

 Scoring of symptoms for AR

Neuropeptides induce vasodilation, thus causing congestion (Howarth, 1997). Cytokine secretion upregulates the expression of adhesion molecules on the vascular endothelial cells, thereby enhancing the activation and adhesion of inflammatory cells. An increase in interleukin-4 (IL-4), IL-5, and granulocyte-macrophage colony-stimulating factor (GM-CSF) is associated with a mucosal eosinophilia (Quraishi, et al., 2004).

#### **3. Traditional management of allergic rhinitis**

Effective allergen avoidance can lead to substantial relief of symptoms. However, patients are still not able to avoid their confirmed allergens such as mites or atmospheric pollens under many circumstances. Medication to manipulate the release of mediators is the next step in the management of AR. Table 2 summarizes the classes of pharmacological therapies for the treatment of AR. The two major classes of medication are oral H1 antihistamines and intranasal corticosteroids. According to the guidelines, oral antihistamines are the first-line therapy which relieve sneezing and rhinorrhea (Bousquet, et al., 2001; van Cauwenberge, et al., 2000; Dykewicz, et al., 1998). Topical administration can further minimize systemic adverse effects of antihistamines. H1 antihistamines are often combined with a decongestant to reach sufficient efficacy for nasal congestion (Quraishi, et al., 2004). The new generation of antihistamines acts as inverse agonists that stabilize the inactive conformation of the receptors and reverses constitutive activity of receptors (Oppenheimer & Casale, 2002). Intranasal corticosteroids are recommended as first-line treatment for moderate and severe AR, which are effective in relieving symptoms such as sneezing, rhinorrhea, itching and congestion (Weiner, et al., 1998). Corticosteroids target the inflammatory mechanisms; therefore the amount of oral corticosteroids for long-term treatment should be carefully adjusted to avoid adverse effects such as osteoporosis and growth inhibition in children (Wilson, et al., 1998).

Decongestants can reduce nasal obstruction and congestion by their vasoconstrictive action on α-adrenergic receptors. The application of intranasal decongestants may cause rhinitis medicamentosa. The adverse effects of oral decongestants include elevated blood pressure, tremor, tachycardia, loss of appetite, sleep disturbance. The long term use of decongestant is not recommended. (Quraishi, et al., 2004). Anticholinergics act as muscarinic receptor blocker which inhibits mucus secretion and subsequent rhinorrhea. Through inhibiting degranulation and neosynthesis of inflammatory mediators, mast cell stabilizers are shown to be effective in reducing symptoms of early inflammatory phase and are useful for preventive purposes (Al Suleimani & Walker, 2007). Leukotrienes inhibitors significantly reduce nasal blockade by inhibiting leukotriene synthesis or serving as antagonists for its receptors. Immunotherapy has the potential to provide a permanent cure for the disease. The proposed mechanisms for immunotherapy include suppression of IgE elevation, decrease in neutrophil and eosionophil activity, reduction in mast cell number, inhibition of T-lymphocyte proliferation (Jayasekera, et al., 2007; Pipet, et al., 2009). However, the technique is burdensome which requiring a lengthy series of injection and it may not be applicable to all patients (Naclerio, et al., 2002). Although most of the drugs are effective in treating certain symptoms of AR, they all have limitations due to their adverse effects. Due to the complex mechanisms involved in the AR, the ideal treatment for this disease has yet to be discovered.

Phototherapy for the Treatment of Allergic Rhinitis 187

Electromagnetic radiation comprises of radio, microwave, infrared (IR), visible light, UV, Xray and gamma radiation. The phenomenon of light absorption to produce electronic excitation of atoms and molecules has long been accepted by photochemists and photobiologists. In phototherapy, wavelengths used include UV (100-400 nm), visible light

Photobiological reactions often involve the absorption of a specific wavelength of light by the functioning photoacceptor molecule. Photochemical effect is elicited using UV and visible light irradiation, whereas thermal effect is induced with FIR irradiation. Light in the ultraviolet range is absorbed by the protein part of the molecule and the visible and NIR wavelengths are absorbed by the metals. Further analysis of action spectra, it is suggested that the primary photoacceptor for the red-NIR wavelengths in mammalian cells is cytochrome *c* oxidase in terminal respiratory chain (Karu, et al., 2005, 2008). Flavoproteins such as NADH dehydrogenase in the beginning of the respiratory chain is believed to be the photoacceptor for the violet-to-blue spectral range (Karu, 2003). In addition, light induces a wave-like alternating electric field in a medium that is able to interact with polar structures and produce dipole transitions. These dipole transitions may lead to the primary actions at

Early hypothesis of the mechanism of primary action upon visible light irradiation can be divided into two categories: (1) singlet oxygen hypothesis based on the singlet oxygen generation from the endogenous molecules possessing the properties of photosensitizers, such as porphyrins and flavoproteins, upon irradiation (Vladimirov, et al., 2004), and (2) the oxidation-reduction hypothesis based on the excitation in chromophores of cytochromeoxidase complex such as CuA, CuB or heme *a*(*a3*), thereby enhancing the electron transfer rate (Lubart, et al., 2005). Later, the nitric oxide (NO) hypothesis was proposed suggesting that the activity of cytochrome *c* oxidase can be regulated by NO, here light irradiation can reverse the partial inhibition by NO (Karu, et al., 2005). Superoxide anion hypothesis was also suggested because of increased production of superoxide anion by irradiation, possibly through promoting the mitochondrial respiratory chain (Karu, 2003). The transient local heating hypothesis suggested that the irradiation energy may lead to a local transient increase in the temperature of absorbing chromophores, which may cause structural changes and trigger biochemical activity (Hallen, et al., 1993). The primary reactions upon light irradiation mainly occur in the mitochondria, which may lead to the secondary reactions occurring in the nucleus and cytoplasm. The secondary reactions involves cellular signaling cascade including increased intracellular ATP level, activation of transcription factors such as Nuclear factor-kappa B (NF-κB) and AP-1, activation of NADPH oxidase, change of the cellular redox potential to more oxidized direction (increased ROS production), manipulation of Ca2+ concentration, alteration of mitochondrial transmembrane potential (ΔΨm), regulation of inducible nitric oxide synthase (iNOS) activity and intracellular pH, and increased DNA/RNA synthesis (Karu, 2003). Other phenomena such as suppression of inflammatory cytokines, up-regulation of growth factor production, modification of extracellular matrix components, inhibition of apoptosis, stimulation of mast cell degranulation, and up-regulation of heat shock protein are also

IR are invisible electromagnetic waves which are subdivided into three categories: nearinfrared (NIR) (0.8-1.5 μm), middle-infrared (1.5-5.6 μm) and far-infrared (FIR) (5.6-1000

**4. Photobiology**

(400-800 nm) and IR (800-105 nm).

observed (Lin, et al., 2010).

cellular and biochemical levels (Amat, et al., 2006).

Table 2. Current therapeutic agents in use for AR

#### **4. Photobiology**

186 Allergic Rhinitis

Antagonists or inverse agonists for histamine at the histamine receptor

Bind to glucocorticoid receptors, affecting the production of various mediators

Stimulate αadrenergic receptors to induce vasoconstriction

Prevent mast cell degranulation and neosynthesis of inflammatory mediators

Leukotriene synthesis inhibitors, antagonists for the leukotriene receptors

Suppress IgE elevation, neutrophil and eosionophil activity, mast cell number, Tlymphocyte proliferation

Mechanism of action Symptom

relief Adverse effects

First-generation: sedation, impaired mental performance, dry mouth, dry eyes, urinary retention

Intranasal: nose irritation, bleeding Oral: long-term use may cause growth inhibition in children and osteoporosis

Intranasal: rhinitis medicamentosa Oral: elevated blood pressure, tremor, tachycardia, loss of appetite, sleep disturbance

Minimal

Possibility of neuropsychiatric side effects

Unknown effects from the modification of immune system

Sneezing, rhinorrhea, itching

Sneezing, rhinorrhea, itching, congestion

Congestion

blockade Rhinorrhea Minimal

Sneezing, rhinorrhea, itching, congestion

Sneezing, rhinorrhea, congestion

Early or late inflammatory phase responses

Class

Antihistamines

Corticosteroids

Anticholinerg

Mast cell

Leukotriene

Immunotherapy

Decongestants Intranasal,

stabilizers Intranasal

inhibitors Oral

Subcutaneous injection

Table 2. Current therapeutic agents in use for AR

Route of administrati -on

Intranasal, oral

Intranasal, oral

oral

Electromagnetic radiation comprises of radio, microwave, infrared (IR), visible light, UV, Xray and gamma radiation. The phenomenon of light absorption to produce electronic excitation of atoms and molecules has long been accepted by photochemists and photobiologists. In phototherapy, wavelengths used include UV (100-400 nm), visible light (400-800 nm) and IR (800-105 nm).

Photobiological reactions often involve the absorption of a specific wavelength of light by the functioning photoacceptor molecule. Photochemical effect is elicited using UV and visible light irradiation, whereas thermal effect is induced with FIR irradiation. Light in the ultraviolet range is absorbed by the protein part of the molecule and the visible and NIR wavelengths are absorbed by the metals. Further analysis of action spectra, it is suggested that the primary photoacceptor for the red-NIR wavelengths in mammalian cells is cytochrome *c* oxidase in terminal respiratory chain (Karu, et al., 2005, 2008). Flavoproteins such as NADH dehydrogenase in the beginning of the respiratory chain is believed to be the photoacceptor for the violet-to-blue spectral range (Karu, 2003). In addition, light induces a wave-like alternating electric field in a medium that is able to interact with polar structures and produce dipole transitions. These dipole transitions may lead to the primary actions at cellular and biochemical levels (Amat, et al., 2006).

Early hypothesis of the mechanism of primary action upon visible light irradiation can be divided into two categories: (1) singlet oxygen hypothesis based on the singlet oxygen generation from the endogenous molecules possessing the properties of photosensitizers, such as porphyrins and flavoproteins, upon irradiation (Vladimirov, et al., 2004), and (2) the oxidation-reduction hypothesis based on the excitation in chromophores of cytochromeoxidase complex such as CuA, CuB or heme *a*(*a3*), thereby enhancing the electron transfer rate (Lubart, et al., 2005). Later, the nitric oxide (NO) hypothesis was proposed suggesting that the activity of cytochrome *c* oxidase can be regulated by NO, here light irradiation can reverse the partial inhibition by NO (Karu, et al., 2005). Superoxide anion hypothesis was also suggested because of increased production of superoxide anion by irradiation, possibly through promoting the mitochondrial respiratory chain (Karu, 2003). The transient local heating hypothesis suggested that the irradiation energy may lead to a local transient increase in the temperature of absorbing chromophores, which may cause structural changes and trigger biochemical activity (Hallen, et al., 1993). The primary reactions upon light irradiation mainly occur in the mitochondria, which may lead to the secondary reactions occurring in the nucleus and cytoplasm. The secondary reactions involves cellular signaling cascade including increased intracellular ATP level, activation of transcription factors such as Nuclear factor-kappa B (NF-κB) and AP-1, activation of NADPH oxidase, change of the cellular redox potential to more oxidized direction (increased ROS production), manipulation of Ca2+ concentration, alteration of mitochondrial transmembrane potential (ΔΨm), regulation of inducible nitric oxide synthase (iNOS) activity and intracellular pH, and increased DNA/RNA synthesis (Karu, 2003). Other phenomena such as suppression of inflammatory cytokines, up-regulation of growth factor production, modification of extracellular matrix components, inhibition of apoptosis, stimulation of mast cell degranulation, and up-regulation of heat shock protein are also observed (Lin, et al., 2010).

IR are invisible electromagnetic waves which are subdivided into three categories: nearinfrared (NIR) (0.8-1.5 μm), middle-infrared (1.5-5.6 μm) and far-infrared (FIR) (5.6-1000

Phototherapy for the Treatment of Allergic Rhinitis 189

Rhinoconjunctivitis Quality of Life Questionnaire (Cingi, et al., 2009). Another prospective, randomized, single-blind study showed that total nasal scores decreased in both mUV/VIS and low-intensity visible light (mUV/VIS without UV) treated groups (Cingi, et al., 2010). But the decrease was highly significant in the mUV/VIS treated group when compared with the low-intensity visible light treated group. However, the impact of endonasal phototherapy on the number of Langerhans cells in the nasal mucosa was limited (Brehmer & Schön, 2010). DNA damage was significantly higher in nasal cytology samples collected immediately after the last treatment (Koreck, et al., 2007). The DNA damage induced by intranasal UV phototherapy was efficiently repaired two months after ending therapy. One of the possible mechanisms that explain the immunosuppressive effect of mUV/VIS is the induction of apoptosis of T cells and eosinophils after UV damage, thus, leading to the inhibition of synthesis and release of pro-inflammatory mediators (Kemény & Koreck, 2007). The side effect of phototherapy is dryness of nasal mucosa, which can be overcome with emollients. Another disadvantage of UV-B treatment is the risk of carcinogenesis. Therefore, it is important to develop phototherapeutic devices using wavelengths other than UV

Very few papers report the application of the light in the other wavelengths in addition to UV light for the management of AR despite that over 2500 papers have been published regarding low level light therapy as a therapeutic modality to speed up tissue repair as well as related biochemical, cellular, histological and functional effects. NIR irradiation suppressed contact hypersensitivity reaction in rats via systemic immunomodulatory effect (Kandolf-Sekulovic, et al., 2003). A double-blind randomized study showed that 70% improvement of clinical symptoms on AR after intranasal illumination at 660 nm (Neuman & Finkelstein, 1997). Thus, light in red or NIR wavelengths with different mechanism of action from UV and visible light may be an ideal candidate for the intervention of AR after

FIR therapy, a non-invasive and convenient therapeutic modality, can improve blood flow and inflammatory status through both its thermal and non-thermal effects. By applying FIR therapy to the nasal region in the patients with AR, our study demonstrated that FIR therapy could improve significantly for the clinical symptoms of eye itching, nasal itching, nasal stuffiness, rhinorrhea and sneezing during period of therapy (Hu & Li,

Thirty-one patients with perennial AR enrolled in the study completed the FIR therapy. All patients had daily symptoms despite antihistamines and local steroid spray treatments. Patients with severe deviation of the nasal septum causing bilateral nasal obstruction and suffering from sinusitis were excluded from the study. A FIR emitter was used for FIR therapy in this study. The wavelength of the light generated from the electrified ceramic plates of this emitter was in the range between 5 and 12 μm with a peak at 8.2 μm. The radiator was positioned via facing patient's nasal region at a distance of 30 cm. The therapeutic time was 40 minutes everyday for 7 days. All the FIR therapies were performed in the morning between 9 am and noon. During the course of the study, the patients did not receive any other anti-allergic management. The effects of FIR on the clinical symptoms

(Morita, et al., 2008).

systematic study.

2007).

**5.1 Far infrared irradiation in allergic rhinitis**

were analyzed by the paired sample t-test.

μm). Different photobiological mechanisms are involved when FIR radiation is employed in phototherapy. The main principles of FIR are radiation, deep penetration and absorption of resonance. At molecular level, FIR exerts rotational and vibrational effects that are biologically beneficial. FIR therapy is often used as alternative physical therapy to decrease joint stiffness, relieve muscle spasms, assist soft tissue injury repair, lead to pain relief, and help to resolve inflammatory infiltrated edema. FIR therapy can improve blood flow and survival of the arteriovenous fistula in hemodialysis patients (Lin, et al., 2007). Furthermore, FIR stimulation on acupoints at Qihai, Kuan yuan and Chung chi decreases both stress and fatigue levels as well as stimulates autonomic nervous system activity in hemodialysis patients (Su, et al., 2009). IR radiation is believed to transfer energy that is perceived as heat by thermoreceptors in the surrounding skin (Inoué & Kabaya, 1989). The abdominal skin temperature steadily increased to a plateau between 38 and 39oC when the top FIR radiator was 20 cm above the rats (Yu, et al., 2006). The expression of HSP70 participates in cytoprotection and may be induced by hyperthermia, infection, UV radiation, NO, etc. However, an *in vitro* study demonstrated that FIR radiation inhibited the proliferation of cancer cells by the low expression level of heat shock protein 70A (Ishibashi, et al., 2008). In addition to the thermal effect, the non-thermal biological effects of FIR therapy led to enhance the extensibility of collagen tissue, stimulate the secretion of transforming growth factor-β1, and increase microcirculation via L-arginine/NO pathway (Toyokawa, et al., 2003; Yu, et al., 2006). Repeated FIR therapy could upregulate the expression of endothelial NO synthase (eNOS) (Akasaki, et al., 2006). FIR therapy was found to exert an antiinflammatory effect via the induction of heme oxygenase-1 in endothelial cells via stimulating NF-E2-related factor (Nrf2) dependent promoter activity. TNF--induced expression of E-selectin, vascular cell adhesion molecule-1 (VCAM-1), and intercellular cell adhesion molecule-1 (ICAM-1) were suppressed (Lin, et al., 2008). A study on the mechanism of action demonstrated that FIR radiation activated p38 and extracellular signalregulated kinase (ERK), but not Akt or c-Jun N-terminal protein kinases (JNK), and significantly promoted angiogenesis by increasing tube formation and the migration of endothelial cells (Rau, et al., 2010). Based on the above studies, the mechanism of action by FIR therapy can be both thermal and non-thermal effects.

#### **5. Phototherapy in allergic rhinitis**

Phototherapy is an effective treatment modality in inflammatory and immue mediated dieases. It has been successfully used in dermatology practice for several decades. The XeCl UV-B laser irradiation and mixed irradiation with UV-A (25%), UV-B (5%) and visible light (70%) (mUV/VIS) resulted in a dose-dependent inhibition of the allergen-induced wheal formation on the skin (Cosma, et al., 2004; Koreck, et al., 2004). Development of new phototherapeutic devices made it possible to treat the inflammatory disease of the nasal mucosa. Intranasal UV-B phototherapy with medium-dose 308 nm XeCl excimer significantly suppressed the nsasl symptoms of patients with severe hay fever (Cosma, et al., 2004). Rhinophototherapy consist of using mUV/VIS resulted in a significant improvement of clinical symptoms for sneezing, rhinorrhea, nasal itching, and total nasal score (Koreck, et al., 2005). The number of eosinophils and the level of eosinophil cationic protein and IL-5 were also reduced. Statiscally significant differences were found in the average results of the