many people , approximately 20% of the population , suffer from allergic rhinitis worldwide . from the clinical point of view ,
allergic rhinitis causes sneezing , nasal discharge , and nasal obstruction . from the immunological point of view ,
allergic rhinitis is a typical th2 immune disorder characterized by a high level of antigen specific ige production . because the enhanced ige production and inflammatory response in rhinitis are due to predominant production of th2 cytokines such as il-4 , the allergic symptoms can be alleviated by inhibition of th2 cytokine responses .
many researchers have investigated allergic rhinitis using an animal model.1 , 2 , 3 the model of allergic mice made by ova sensitization and challenge had increased serum ige and eosinophil .
shin'iseihaito ( ssht ; magnolia flower lung - clearing decoction ;  xn y qng fi tng ) , a formula consisting of nine crude drugs , has been used for the treatment of nasal diseases such as chronic sinusitis in traditional japanese kampo medicine (  r bn hn y ) and traditional chinese medicine ( tcm ;  zhng y).4 , 5 more than 50% of individuals with allergic rhinitis have clinical or radiographic evidence of chronic sinusitis and 2558% of individuals with sinusitis have aeroallergen sensitization .
previous researches suggest that chronic sinusitis could be an atopic disease driven by ige sensitization to aeroallergens . from these studies
however , it has not been clarified for its role in anti - allergy therapy before .
thus , in the present study , we investigated whether ssht is able to suppress the murine allergic reaction induced by nasal sensitization .
female balb / c mice ( japan slc ltd , hamamatsu , japan ) were used .
shin'iseihaito consists of 3.0  g ( daily dose for human ) of gypsum fibrosum (  sh go ) , 3.0  g of tuber of ophiopogon japonicus (  mi mn dng ) , 1.5  g of root of scutellaria baicalensis (  hung qn ) , 1.5  g of rhizome of anemarrhena asphodeloides (  zh m ) , 0.75  g of fruit of gardenia jasminoides (  zh z ) , 1.5  g of bulb of lilium lancifolium (  bi h ) , 1.5  g of flower of magnolia salicifolia (  xn y ) , 0.5  g of leaf of eriobotrya japonica (  p p y ) , 0.75  g of rhizome of cimicifuga heracleifolia (  shng m ) .
these crude drugs were boiled , filtered , and the decoction was dried to yield powdered extract ( ssht , 2.5  g for daily human dose ) .
ssht ( lot : 14b019 ) was provided as a generous gift from the kobayashi pharmaceutical co. , ltd ( osaka , japan ) .
ssht was suspended in distilled water to prepare the stock solution at a concentration of 0.1  g / ml and kept in  20  c until use .
allergic murine models were established based on the previously described methods with minor modifications.7 , 8 the experimental timetable is provided in fig . 
1 . mice were intraperitoneally administered 0.1  mg / ml ova ( sigma  aldrich , st . louis , mo , usa ) and 40  mg / ml al(oh3 ) in saline at a dosage of 100  l / mouse .
sensitization was repeated twice ( days 0 and 7 ) , followed by daily injections of ova solution ( 15  mg / ml in saline , 10  l / each nostril ) into nostrils from day 1428 ( challenge ) . in the ssht - treated group , mice were force - fed ssht ( 10  mg/0.1  ml/10  g body weight ( bw)/day , 20-fold of human dosage ) from day 1328 .
mice in the control group were given an equal volume of saline and were infected using the same method . at the end of animal experiment ,
all animal procedures were approved by the institutional animal care and use committee at nagoya city university , japan .
blood samples from the allergic murine model were collected after 2  h of the last challenge on day 28 .
the number of leukocyte and eosinophilia were measured in tohkai cytopathology institute ( gifu , japan ) .
concentrations of serum total ige , il-4 , and ifn- were evaluated using mouse ige elisa , mouse il-4 , and mouse ifn- elisa kit ( biolegend inc .
san diego , ca , usa ) according to the manufacturer 's instructions , respectively .
cytokine levels were calculated using standard murine recombinant cytokine curves run on the same immunoplate .
the untreated female 6 week - old balb / c mice were injected intradermally with 10  l aliquot of 50 fold diluted anti - ova of serum in saline into shaved dorsal skin sites .
after two days , ova ( 0.1  mg ) with 0.5% evans blue ( wako pure chemicals , osaka , japan ) in saline was injected intravenously into the tail vein .
one hour after antigen challenge , mice were euthanized and the dorsal skin of the mouse was removed to measure the pigment area .
the area of blue spots on the internal surface of the skin was measured.10 , 11 statistical analysis was performed by repeated one - way analysis of variance ( anova ) and the tukey / bonferroni / dunnett 's multiple comparison test .
a probability value ( p  <  0.05 ) was considered to be statistically significant .
the leukocyte levels were significantly higher in allergic mice than those in untreated mice , but those levels were not significantly changed by ssht - treatment [ fig . 
the eosinophilia levels were significantly higher in allergic mice than those in untreated mice . compared to allergic mice ,
the allergic rhinitis mice treated with ssht had significantly decreased eosinophil levels ( p  <  0.01 ) [ fig . 
the ige levels were higher in allergic mice than those in untreated mice . compared to allergic mice ,
the il-4 levels were significantly higher in allergic mice than those in untreated mice . compared to allergic mice ,
the allergic mice treated with ssht had also significantly decreased il-4 levels ( p  <  0.01 ) [ fig . 
the ifn- levels were significantly lower in allergic mice than those in untreated mice . compared to allergic mice ,
the allergic mice treated with ssht had also significantly increased ifn- levels ( p  <  0.01 ) [ fig . 
3c ] . as we found that the eosinophilia and cytokine level of allergic mouse treated with ssht were remarkably decreased
the sera containing ova - specific anaphylactic antibodies were intradermally injected and the pca reaction was measured 1  h after the injection of the evans blue solution containing ova .
we observed that pca reactions using the sera of allergic mice were exhibited as blue spots .
however , the average sizes of blue spots were significantly decreased in the sera of ssht - treated allergic mice compared to those in the sera of allergic mice .
in the present study , we showed that the administration of shin'iseihaito ( ssht ; magnolia flower lung - clearing decoction ;  xn y qng fi tng ) is capable of suppressing the ige and il-4 levels in murine allergic reaction induced by nasal sensitization .
thus , our result showed that ssht may return the decreased ifn-/il-4 ( th1/th2 ) ratio of the allergic group to the normal range .
this result may also explain the reasons why the administration of ssht suppressed the production of anti - ova ige antibody without affecting the development of cd4 t cells , since il-4 is the cytokine known to induce immunoglobulin class switching to ige .
il-4 is known to serve not only as a mast cell growth factor but also as the major mast cell chemoattachment .
therefore , it is possible that the decrease of th2 responses in nasal mucosa modulated the mast cell ( ige)-mediated nasal symptoms , in concert with the decreased production of anti - ova ige antibody .
our result of pca reaction also demonstrated that ssht inhibited the anaphylaxic factors such as ige and other antigens .
the precise mechanism of the effect of ssht on pca reaction is to be determined in a future study . although our results demonstrate that ssht had anti - inflammatory effect , each individual component of ssht also shows anti - inflammatory effect .
especially , the flower of m. salicifolia (  xn y ) has several anti - inflammatory effects including the inhibitory effects of mast cell - derived histamine release and pca reaction.14 , 15 , 16 , 17 the fruit of g. jasminoides (  zh z ) inhibits the histamine release from mast cells and lowered the serum level of ige and histamine in allergic murine model .
the rhizome of a. asphodeloides (  zh m ) inhibits nf-b transcription activity via the p38 mapk and erk pathway .
the leaf of e. japonica (  p p y ) had anti - inflammatory effect with attenuation of p38map kinase and erk .
some researcher reported that the root of s. baicalensis (  hung qn ) , the bulb of l. lancifolium (  bi h ) , and the tuber of o. japonicus (  mi mn dng ) had anti - inflammatory activity.21 , 22 , 23 although each component has anti - inflammation effect , this mechanism in detail has been unknown .
japanese traditional kampo medicine (  r bn hn y ) is generally composed of several components and the interaction of them may enhance the effect of drugs . as further investigation from this perspective is needed , ssht may have pronounced anti - inflammatory effects .
administration of shin'iseihaito ( ssht ; magnolia flower lung - clearing decoction ;  xn y qng fi tng ) is capable of improving allergic status in ova - induced allergic murine model .
further studies are required to confirm the anti - allergy effects and elucidate the mechanisms of anti - allergy action of ssht in allergic murine model .
 shin'iseihaito ( magnolia flower lung - clearing decoction ;  xn y qng fi tng ) , a formula of traditional japanese kampo medicine (  r bn hn y ) and traditional chinese medicine ( tcm ;  zhng y ) , has been used for the treatment of chronic sinusitis . 
 the objective of this study was to evaluate the anti - allergic effect of shin'iseihaito on murine allergic reaction induced by nasal sensitization using ovalbumin ( ova ) as an antigen . 
 extract of shin'iseihaito ( ssht ) could reduce the eosinophil , serum ige and interleukin ( il)-4 levels , while increased the interferon ( ifn)- levels in allergic mouse . 
 furthermore , allergic - murine serum treated with ssht could not activate passive cutaneous anaphylaxis ( pca ) reaction in murine model . 
 thus , our study showed that ssht may possess anti - allergic activity . 
 we suggested that ssht may contribute to inhibit the exacerbation of allergic reaction induced by nasal sensitization . 