the prevalence of asthma is approximately 300 million cases in the world and in thailand it affects approximately 6.9% of adults . the airway becomes swollen and inflamed and the muscles around the airway contract causing the bronchial tubes to narrow . symptoms and exacerbations of asthma include breathlessness , coughing , wheezing , and chest tightness . severe exacerbations of asthma are potentially life - threatening and require prompt care , close observation for deterioration , and frequent treatments . the public health burden of acute exacerbation asthma is an especially important and costly problem . vitamin d is a fat - soluble vitamin that is important to the body by balancing calcium and bone , innate and adaptive immunity , and homeostasis of many organs . vitamin d from food or dermal synthesis is inactive and requires enzymatic conversion to become active . hypovitaminosis d is typically diagnosed by measuring the concentration in blood of 25-hydroxyvitamin d ( 25(oh)d ) , the primary circulating form of vitamin d , and then 1,25-dihydroxyvitamin d , the active form of vitamin d , by the enzyme 25-hydroxylase in the liver and kidney . vitamin d insufficiency is defined as a 25(oh)d level less than 30  ng / ml . furthermore , vitamin d deficiency is defined as 25(oh)d levels below 20  ng / ml , with a resultant consistent elevation of pth and reduction in intestinal calcium absorption . people living in tropical countries are exposed to the sun and usually have high levels of vitamin d. however , studies in tropical countries have found the prevalence of vitamin d deficiency to be about 30 to 50% [ 8 , 9 ] . in thailand , low serum levels of vitamin d have been linked to increased risk of asthma exacerbation in children and adults . hypovitaminosis d is associated with increased risk of cardiovascular disease , autoimmune disease , cancer , and allergy disease [ 11 , 12 ] . it directly affects the adaptive immune system through its effects on th1 , th2 , and regulatory t cells . it promotes peripheral tolerance by inhibiting inflammation and the induction or maintenance of regulatory t cell populations , both il-10 and foxp3 . this cross - sectional study demonstrated that the frequency of circulating cd4foxp3 treg cells is significantly lower in steroid resistance than in steroid sensitive asthmatic patients . also , it directs the induction of innate antimicrobial mechanisms to efficiently resolve infection , especially respiratory viral infections . notably , vitamin d insufficiency has been associated with severe asthma exacerbations in children and decreased serum vitamin d levels are associated with increased corticosteroid use [ 18 , 19 ] . vitamin d deficiency has been associated with the asthma epidemic , obesity in african americans , westernization of countries with higher - risk populations for asthma , increased airway hyperresponsiveness , lower pulmonary functions , worse asthma control , and possibly steroid resistance . the average number of asthma attacks per person per year decreased from 0.44 to 0.28 with vitamin d . vitamin d reduced the risk of attending hospital with acute exacerbation asthma from 6 per 100 to around 3 per 100 . however , in tropical countries like thailand , data on the association between serum vitamin d level and asthma exacerbation in adult patients is limited . this study compared serum vitamin d levels during the period of asthma exacerbation and during the periods in stable condition . also , patients with low vitamin d levels received vitamin d2 supplement for three months and then asthma control and lung function were monitored . adults with asthma exacerbation were recruited from the allergy clinic and emergency department , phramongkutklao hospital , bangkok , thailand . severe asthma exacerbation is defined by acute episodes of progressively worsening shortness of breath , coughing , wheezing , and chest tightness . a patient with asthma exacerbation requires treatment with nebulized short - acting beta-2 agonist and oral corticosteroid added to daily asthma medications . we enrolled patients ( 18 years of age or older ) with asthma exacerbation and without upper respiratory tract infection . serum 25(oh)d , asthma control test ( act ) score , and peak expiratory flow rate ( pefr ) were measured during the period of severe exacerbation . at least two weeks after the exacerbation , the patients during the period in stable condition ( control asthma without inhaled short - acting beta-2 agonist ) visited the allergy clinic again to check serum vitamin d levels , act score , and pefr . a clinical history of asthma was confirmed by bronchodilator responsiveness ( as defined by an improvement in the fev1 of > 200  ml and 12% after administration of four puffs ( 360  mg ) of salbutamol by metered dose inhaler ) . participants with significant medical illness other than asthma , those with a history of respiratory tract infection in the past seven days , those with nonadherence to use control medication , and those taking vitamin d supplement were excluded . the control subjects were adults without a history of chronic illness . during the period of control asthma , if the serum level of 25(oh)d was less than 30  ng / ml , the subjects were treated with the ergocalciferol ( vitamin d2 ) supplement at a dose of 20,000  iu every alternate day . after 3 months , subjects were evaluated again for serum 25(oh)d , act score , and pefr . during 3 months of vitamin d2 supplement , subjects received the same control asthma medication , that is , inhaled long - acting beta-2 agonist plus corticosteroid and leukotriene receptor antagonists . the study was approved by the institutional review board , royal thai army medical department . pefr was measured during forceful exhalation , starting from full lung inflation with the patient standing . serum total 25(oh)d was quantified by high - performance liquid chromatography - tandem mass spectrometry and categorized in 25(oh)d sufficiency  30  ng / ml , insufficiency < 30  ng / ml , and deficiency < 20  ng / ml . therefore , the serum level of 25(oh)d was considered the best biomarker of vitamin d metabolic status and reflected contributions from all sources of vitamin d. patients were divided into three groups : complete asthma control ( act = 25 ) , partial asthma control ( act 2124 ) , and poor asthma control ( act < 20 ) . data were analyzed using graphpad prism 6 ( graphpad software , la jolla , ca , usa ) . data description was primarily based on means and standard deviations ( sd ) for continuous endpoints and on frequencies for categorical parameters . nonparametric comparisons between asthma exacerbation and asthma control were made using wilcoxon matched - pairs signed test . the mann whitney test is a method to compare two population means that come from the same population . we assessed the association between serum vitamin d levels and act that was evaluated by spearman 's correlation . a p value < 0.05 was considered statistically significant . a total of 47 adult patients with exacerbation asthma were enrolled and baseline characteristics are shown in table 1 . a total of 13 of 16 ( 81.25% ) patients with age > 70 years had abnormal vitamin d levels . of these , three patients had a vitamin d level < 10  ng / ml . subjects were predominantly female ( 74.47% ) . also , females had more abnormal d levels ( 82.85% ) than males ( 41.66% ) . thirty - four subjects ( 72.34% ) with asthma exacerbation had vitamin d levels below normal . therefore , patients that had uncontrolled asthma ( act < 20 ) and vitamin d deficiency constituted a third of the total . in the healthy control group , females comprised 65% and the mean age was 63  17.29 years . the prevalence of vitamin d insufficiency in the healthy control group was 47.5% without vitamin d deficiency . mean 25(oh)d concentrations were 23.84  8.89  ng / ml in the period of asthma exacerbation and increased to 24.34  9.8  ng / ml in terms of no exacerbation but differed insignificantly , p = 0.6 ( figure 1(a ) ) . in the healthy control group , mean 25(oh)d concentration was 31.75  7.28  ng / ml . to compare between asthmatic patients and healthy controls , the mean 25(oh)d levels in the period with asthma exacerbation and without exacerbation in the asthma group were significantly lower than those in the healthy control group , p = 0.0002 and p < 0.0001 , respectively ( figure 1(a ) ) . analyzing mean 25(oh)d concentrations between females , 22.55  7.74 ng / ml , and males , 27.59  11.16  ng / ml , revealed no significant difference ( figure 1(b ) ) . however , females had a higher risk of developing vitamin d deficiency ( or = 5.83 ; 95% ci = 1.1230.4 , p = 0.03 ) and vitamin insufficiency than males ( or = 8.17 ; 95% ci = 1.3051.4 , p = 0.02 ) . severe asthma exacerbation mean 25(oh)d concentrations , found between ages 50 , 5159 , 6069 , and 70 years , were 33.57  5.77  ng / ml , 23.67  6.68  ng / ml , 23.2  7.4 , and 20.24  9.82 , respectively . age 50 years presented more significantly higher vitamin d levels than other age groups ( figure 1(c ) ) . also , in the period without exacerbation , age 50 had higher vitamin d levels than 70 years ( figure 1(c ) ) . no significant difference was observed between partly controlled ( act < 20 ) and uncontrolled asthma ( act 2124 ) in mean 25(oh)d concentration with asthma exacerbation and without asthma exacerbation ( figure 1(d ) ) . for asthmatic patients with % predicted pefr > 80% , mean 25(oh)d concentration 26.48  6.17  ng / ml was significantly higher than in asthmatic patients with % predicted pefr < 60% ( 18.81  7.04  ng / ml , p = 0.02 ) ( figure 1(e ) ) . in patients with % predicted pefr > 80% , mean 25(oh)d concentrations between with asthma exacerbation 26.48  6.17 and without asthma exacerbation 24.34  6.36  ng / ml significantly differed , p = 0.009 ( figure 1(e ) ) . representative vitamin d status and asthma control test score are shown in figure 2(a ) . patients with sufficient vitamin d levels had significantly higher act scores than patients with vitamin d deficiency during asthma exacerbation ( 20.23  1.83 versus 15.11  5.73 , p = 0.04 ) ( figure 2(a ) ) . moreover , during the period without asthma exacerbation , patients with vitamin d insufficiency and deficiency significantly increased asthma control test score to compare in the period with exacerbation asthma ( figure 2(a ) ) . vitamin d status presented no difference in % predicted pefr ( figure 2(b ) ) . we next tested whether serum vitamin d levels were associated with age , controlled asthma , and % predicted pefr . increasing age inversely correlated with serum vitamin d level ( p = 0.003 , r = 0.30 ) ( figure 3(a ) ) . then serum 25(oh)d levels significantly positively correlated with controlled asthma ( p = 0.006 ; r = 0.30 ) ( figure 3(b ) ) . however , an association between serum vitamin d levels and lung function was not observed ( figure 3(c ) ) . twenty - four of 34 subjects ( 70.58% ) who had serum 25(oh)d level less than 30  ng / ml participated in receiving vitamin d2 supplement ( 12 subjects denied treatment with vitamin d supplement ) . we prescribed a vitamin d2 supplement , ergocalciferol ( vitamin d2 ) , 20,000  iu every alternate day for 3 months to 24 asthmatic patients ( 7 , partly controlled , and 17 , uncontrolled asthma ) . after 3 months of vitamin d administration , mean 25(oh)d concentration was significantly higher than baseline among asthmatic patients ( 20.41  5.13 versus 36.41  5.22  ng / ml , p < 0.0001 ) ( figure 4(a ) ) . to compare with baseline , in the uncontrolled asthma group , mean act score at baseline , 16.12  1.03 , significantly increased after receiving vitamin d supplement for 3 months ( 20.37  3.55 , p = 0.0004 ) ( figure 4(b ) ) . the lung function in the partly controlled and uncontrolled asthma group showed slightly increased % predicted pefr after 3 months of vitamin d supplement but without significant difference ( figure 4(c ) ) . next , we observed % change of act score and percent predicted pefr between partly controlled and uncontrolled asthma group . uncontrolled asthma group had significantly improved act score more than partly controlled asthma group ( figure 4(d ) ) . no significant difference with % change of percent predicted pefr between both groups was observed ( figure 4(e ) ) . this cross - sectional study examined vitamin d levels in adult asthmatic patients with differing periods of severe asthma exacerbation and without exacerbation . the study could not conclude that decreased vitamin d level was the leading cause of severe asthma exacerbation among asthmatic thai adults . however , many studies have suggested vitamin d deficiency was linked to increased risk of asthma exacerbation . vitamin d insufficiency among north american and puerto rican children was associated with severe asthma exacerbations [ 25 , 26 ] . moreover , adults presenting vitamin d deficiency were reported in to be linked to the risk of severe asthma exacerbations . furthermore , a large cohort from israel showed asthmatic patients with vitamin d deficiency had a 25% higher chance of having an exacerbation than patients within normal range . however , many studies examined serum vitamin d level only one time and did not compare with the period without exacerbation in the same patient . we showed vitamin d insufficiency and deficiency were more highly prevalent among asthmatic patients than among healthy people . similarly , the study in asthmatic thai children showed the prevalence of decreased vitamin d status was 64% . thailand , a southeast asian country , is located close to the equator and has a tropical rainforest climate with high sun exposure , one source of vitamin d among humans . one possible explanation is that vitamin d metabolism is influenced by various factors including skin pigmentation , sunscreen use , clothing , drugs , smoking , age , obesity , and several chronic illnesses . dietary intake and supplements are a secondary source of vitamin d. few natural vitamin d rich food sources are available in thailand and foods are not fortified with vitamin d . avoiding sun exposure and low intake of vitamin d may play significant roles in vitamin d insufficiency among people . the study by chailurkit et al . among 2,641 thai adult subjects found the prevalence of vitamin d insufficiency was dependent on geographic region and associated with females , younger age , and living in an urban area . the findings of the present study confirmed report that patients with various degrees of vitamin d status were associated with asthma severity , monitored among children and adults [ 27 , 33 ] . asthma has been treated by vitamin d supplement in at least three adult random controlled trials ( rct ) . conducted an rct enrolling 130 patients with moderate persistent asthma and found vitamin d supplementation to be associated with asthma controllers that could significantly improve fev1 in mild to moderate persistent asthma after 24 weeks . in addition , yadav and mittal reported monthly doses of 60,000  iu vitamin d3 significantly reduced the number of exacerbations as compared with placebo and pefr significantly increased in the treatment group . moreover , the study by de groot et al . conducted an rct involving 44 patients with nonatopic asthma and found vitamin d supplementation reduced eosinophilic airway inflammation among patients with nonatopic asthma with severe eosinophilic airway inflammation . studied rct involving 408 adult asthmatic patients and concluded vitamin d3 did not reduce the rate of exacerbation among adults with persistent asthma and vitamin d insufficiency . in addition , martineau et al . conducted an rct comprising 250 adults with asthma in london and reported bolus - dose vitamin d3 supplementation did not influence time to exacerbation in a population of adults with asthma . the more recent rct results suggest likely interactions of the vitamin d acute respiratory tract infection association by baseline vitamin d status ( i.e. , vitamin d deficient individuals receive more benefit than those with normal concentrations ) and possibly greater advantage for subgroups that are at higher risk of acute respiratory tract infection , such as young children , older adults , or individuals with immune defects . providing vitamin d supplements to improve asthma symptoms and the limitation of the study is no randomized control trial and no control group in vitamin d supplement study that it is difficult to conclude the benefit of vitamin d supplement in uncontrolled asthmatic patients . in addition , the subjects in this study are of low number because asthma exacerbation with upper respiratory tract infection is common in thailand ; this study excluded the cause of asthma exacerbation with upper respiratory tract infection . we used vitamin d2 supplementation because the cost of the treatment is less than vitamin d3 supplementation . however , vitamin d2 supplementation is effective in increasing serum 25(oh)d level and not toxic when provided in large amounts . further study will be planned to do randomized control trial and multicenter study to prove the benefit of vitamin d supplement in asthmatic patients . hypovitaminosis d was common in patients with asthma exacerbation , but the low vitamin d level did not cause asthma exacerbation . the study showed the benefits of examining serum 25(oh)d and providing vitamin d supplements to uncontrolled asthmatic patients .