This study, carried out on a large number of subjects, highlights that some seasons may influence the results of methacholine challenge tests. In fact, a lower number of hyper-responsive subjects were found in summer compared with autumn. Furthermore, the logistic regression models showed a higher (+39.3%) bronchial risk of hyper-responsiveness in autumn in comparison with summer when we chose a 2400 μg cut-off value to divide hyper-responsive subjects from normal subjects. There was also a higher risk (+34.7%) for a higher degree of severity in airway hyper-responsiveness (using a cut-off of 400 μg) both in spring and in autumn, again compared with summer. This result is certainly due to a greater allergen exposure in these seasons in comparison with summer. This major exposure may cause an increase in airway inflammation with a consequent increase in BHR. Blood eosinophil numbers were indeed higher in BHR subjects with seasonal allergic rhinitis during pollen season than those without BHR, whereas there was no difference in the number of these cells during off-season in these two groups.[21
] After a nasal challenge with grass pollen in rhinitis, BHR and both eosinophil numbers and eosinophil-cationic protein (ECP) concentration in induced sputum had increased and the latter two were correlated to methacholine responsiveness.[22
] In addition, eosinophils and ECP, both in peripheral blood and sputum, increased in mite-sensitized asthmatics after Dermatophagoides pteronyssinus
exposure and this rise was associated to a PD20
Fruchter and Yigla[14
] , in a similar study, partially confirmed our results as they found a lower number of positive methacholine tests in summer compared with winter and spring, due to a lower allergy burden in mites and in pollens in that season. Another study also showed that the sensitivity of exercise testing for asthma in adolescents is halved in summer due to a decreased asthma activity for a lower exposure to allergens, to air pollutants such as nitrogen oxides and sulfur dioxide and to respiratory viruses such as respiratory syncytial virus and influenza.[24
Other authors have found, in agreement with our study, a greater autumnal airway hyper-responsiveness risk. In fact, some of them had already observed how airway hyper-responsiveness increased in autumn in multiple-sensitized allergic asthmatic patients.[3
] The higher prevalence of BHR in autumn is probably due to an increased exposure to house dust mite in this season. In fact, a more severe airway hyper-responsiveness was found in mite-sensitized patients and this was strongly related to an increase of Der p 1 concentrations in floor dust, particularly at the beginning of autumn.[7
] Other authors have also found that sensitization to house dust mite, as opposed to sensitization to pollen allergens alone, is associated with BHR, thus confirming that mite allergy plays the leading role in BHR.[6
] Unfortunately, in our study, the allergic status of the subjects recruited is not known. However, it seems that 95.4% of young adults, with positive methacholine tests, resulted positive to aeroallergen skin prick tests.[24
] We found a positive skin prick test in approximately 64% of a group of 811 subjects recruited in the same Tuscan area for suspected rhinitis and/or asthma. We found a higher sensitization to mites (61.8%), followed by cypress (56%), grasses (44%), olea (36.4%), and pellitory (19.6%) in these subjects.[26
] Therefore, among the 4 826 subjects considered for our study, a large number of them might be allergic to house dust mites, thus influencing the variations in autumnal airway hyper-responsiveness. However, also atopic asthmatic subjects—who are exposed to high levels of dust mites (as it may happen in autumn) but not sensitized to this allergen—show evidence of increased airway reactivity.[27
The increased mite-induced airway inflammation may also explain the increased obstruction of the airways in autumn demonstrated by a reduced FEV1
/FVC ratio found in this season compared with summer. Consequently, this autumnal increased obstruction may be responsible for the higher prevalence of BHR found in this season. In fact, it is known that the level of BHR is negatively correlated to pulmonary function.[18
The autumnal drop in temperature, with the increase of upper or lower respiratory tract infections, might be another reason for the higher prevalence of hyper-responsiveness in autumn. In fact, some studies correlated bronchial infections with an increased airway hyper-responsiveness.[2
] However, the absence of any BHR increase in winter, when bronchial infections were more frequent, does not explain this increase in hyper-responsiveness in autumn. On the contrary, Fruchter and Yigla found a higher prevalence of BHR in winter and not in autumn.[14
] It is known that in Israel, in particular in Haifa where the study was performed, the climate is usually milder if compared with that of Northern Italy, which makes the Israeli winter similar to the Italian autumn. A very high humidity rate and very cold air may also influence airway hyper-responsiveness,[12
] but humidity and the temperature in autumn do not usually reach extreme values in our area. Therefore, any direct influence of temperature and/or humidity on this BHR variability is unlikely.
Another important result of this study was the observation of a lower PD20
value in spring compared with summer. Furthermore, we found that there was a higher probability of having a PD20
lower than 400 μg in spring compared with the summer. This means that the higher pollen concentrations in spring, due to blooming of grasses, olea, and pellitory in our area, can determine a more severe BHR in subjects sensitized to these pollens. This is confirmed by some studies that have shown an increase in bronchial reactivity in spring in hyper-responsive subjects with rhinitis and/or asthma sensitized to Parietaria, Olea, and Gramineae.[2
] It seems that Parietaria is more important than Olea and Gramineae as a risk factor for non-specific BHR.[28
Our study highlighted that seasonal variability concerns particularly females and overweight/obese subjects. The prevalence of hyper-responsiveness in these subjects was higher in autumn in comparison with what was obtained in the other seasons, which was unexpected. Furthermore, the PD20
value was lower in females and in non-smoker overweight/obese subjects in spring compared with that in summer (eliminating the confounding effect of smoking). We have not found any explanations in literature for this greater prevalence of hyper-responsiveness in autumn and this greater hyper-reactivity in spring, in particular in these categories of subjects. Several studies have shown that the female sex has a higher BHR risk factor[18
] as confirmed also in our study (+53.6% of risk increase). It has been suggested that the higher prevalence of airway hyper-responsiveness in females could be caused by their smaller lung volumes.[27
] On the other hand, there are conflicting studies on a possible existence of a relation between BMI and hyper-responsiveness.[31
] However, we found that an overweight/obese status is an independent high BHR risk (+21.9%). The obesity-related changes in TNF-alpha, leptin, and adiponectin may contribute to this BHR increase.[35
] Therefore, for this particular predisposition to airway hyper-responsiveness, females and obese subjects probably develop a greater sensitivity to house dust mites and pollens compared with other categories of subjects (for example, males or subjects with normal weight). In autumn, when house dust mites increase, and in spring, when the pollen exposure is high, these subjects may show a higher reactivity to these allergens. This may suggest a different asthma phenotype in female and overweight/obese subjects. Children-adolescents also show a lower PD20
in autumn compared with winter. This season variability is probably due to a greater susceptibility to mites or pollen in young subjects with a small airway calibre where the allergic response already tends to be higher itself. Furthermore, in non-smokers aged between 20 and 35 years old (eliminating the confounding effect of smoking), a lower PD20
was found in spring compared with summer, probably due to a stronger bronchial reactivity to pollens in these young subjects rather than in those who were older where this reactivity is known to be lower.[16
This study also confirmed how smoking can be a BHR risk factor (+27.9%) in subjects with typical asthma respiratory symptoms. Other studies had already observed how asymptomatic smokers showed an increased airway responsiveness compared with non-smokers.[36
] Furthermore, allergic rhinitis (+/−asthma) hyper-responsive subjects showed an improvement in methacholine-induced BHR 12 months after they had stopped smoking, thus confirming the importance of such habit in BHR.[38
] Smoking can cause chronic airway neutrophilic inflammation and oxidative epithelium damage, thus leading to the development of airway hyper-responsiveness;[39
] in fact, a dose-dependent relationship between the number of cigarettes smoked and the degree of hyper-responsiveness has been demonstrated.[40
] A large number of non-smokers showed a BHR in autumn compared with what was found in summer, whereas no seasonal variations were observed in smokers. Smoking probably induces a persistent inflammation or the worsening of the pre-existent inflammation induced by asthma which cancels the seasonal changes in BHR over the course of the year. The absence of smoke-induced inflammation makes this airway phlogosis susceptible to allergen-related seasonal changes in non-smokers.
We also analyzed if there were any seasonal variations in the different levels of BHR. We did not find any differences neither in prevalence nor in PD20
among the seasons in moderate-to-severe, mild, and borderline BHR. However, according to the guidelines,[1
] the PC20
cut-off value for a positive methacholine test is 4 mg/ml, comparable with our PD20
dose of 400 μg. Using a cut-off value of 400 μg to define BHR in the logistic regression model, we found a high BHR risk both in autumn and spring. Therefore, subjects with PD20
< 400 μg (with a higher BHR) may show a worsening of BHR in these seasons.
We must also add that the results seem to be in contradiction where the prevalence of BHR was higher in autumn while the PD20 was lower in the spring. This may be due to a higher proportion of subjects with severe BHR in spring (although not significant) compared with summer, but in autumn, we observed that the distribution of subjects with different levels of BHR is the same as that observed in spring. Alternatively, this increased BHR in spring could be due to a greater activity on the airways by spring allergens; on the contrary, autumnal allergens may cause a reduced bronchial reaction in subjects with BHR.
In summary, this study has shown a seasonal variability of airway hyper-responsiveness; therefore, when methacholine tests are repeatedly performed over the course of time (for example, in clinical trials), this must be taken into account. In addition, considering that there is a lower BHR in summer, especially in comparison with spring and autumn, a reduced therapy level in this season may be hypothesized in multi-sensitized persistent asthmatics, whereas in autumn and spring—when their BHR is higher—treatments may be increased.
In conclusion, this study has shown a seasonal variability in the response to the methacholine challenge test in subjects with suggestive asthma symptoms, which is probably due to a different allergens exposure in the various seasons. A higher probability of finding a hyper-responsiveness may be detected in autumn and in spring, whereas a lower one may be found in summer. Spring is the season when BHR may be more severe. This seasonal variability seems to concern in particular females and overweight-obese subjects.