Study On The Clinical Effect And Mechanism Of Wen’s JianPiChuShi Decoction In Treating Chronic Eczema Of Spleen Deficiency And Dampness Encumbrance Type From The Theory Of "Combination Of Clearing And Tonifying Methods"

1.Objective1.1 Summarizing the characteristics of Chinese medicine use in chronic eczema clinical trials in China and the similarities and differences with Sichuan Wen’s school of dermatology based on data mining techniques.1.2 Analysis of the active ingredients,targets and mechanisms of Wen’s Jian Pi Chu Shi Decoction based on network pharmacology technology.1.3 Clinical observation on the efficacy and safety of Wen’s Jian Pi Chu Shi Decoction in treating chronic eczema patients with spleen deficiency and dampness encumbrance.1.4 To explore the effects of Wen’s Jian Pi Chu Shi Decoction and its disassembled formula on the IL-23/IL-17 inflammatory axis and skin barrier by constructing mouse model of chronic eczema.Finally,it will provide experimental basis for the clinical efficacy of Wen’s Jian Pi Chu Shi Decoction and demonstrate the scientific nature of the theory of “combination of clearing and tonifying methods”.2.Materials and methods2.1 The Characteristics of Chinese herbal medicine in chronic eczema clinical trials in China and its similarities and differences with Sichuan Wen’s school of dermatology The Chinese literature on clinical research types of Chinese herbal compounding for chronic eczema was searched in 4 major electronic databases,including CNKI,Wanfang,VIP,and CBM.After manual screening,classification,data entry and other steps,I would summarize the characteristics of Chinese medicine use in clinical trials about chronic eczema and its similarities and differences with Wen’s school of dermatology.2.2 Effective ingredients,targets,and pathways of Wen’s Jian Pi Chu Shi Decoction Using network pharmacology methods to analyze the effective ingredients,targets,and pathways of Wen’s Jian Pi Chu Shi Decoction,and comparing them with loratadine to identify similarities and differences.2.3 Analysis of clinical efficacy and safety of Wen’s Jian Pi Chu Shi Decoction2.3.1 Research objectNinety patients with chronic eczema who met the inclusion criteria of traditional Chinese medicine and western medicine were recruited from the Department of Dermatology,Hospital of Chengdu University of Traditional Chinese Medicine from January 2022 to December2022.They were assigned to the experimental and control groups according to the random grouping principle.2.3.2 Experimental methods and proceduresThe experimental group was given Wen’s Jian Pi Chu Shi Decoction tid/d orally.Patients in the control group were given loratadine tablets 10mg/d orally.The course of treatment was 4 weeks.2.3.3 Observed indicators and Efficacy evaluationEASI score,5-D pruritus score,POEM score,DLQI score,TCM syndrome score and skin barrier function test were performed on all patients enrolled before and after treatment,respectively.In addition,according to the voluntary principle,the changes of IL-23 and IL-17 in serum of patients before and after treatment were examined by ELISA.The efficacy was graded into 4 levels.2.3.4 Safety analysisThe clinical symptoms and laboratory tests(blood routine,liver and kidney function)were used as safety indicators.Recorded and evaluated adverse events.2.4 Study and analysis on the mechanism of Wen’s Jian Pi Chu Shi Decoction2.4.1 Experimental animal grouping,chronic eczema model construction and treatment methodsThirty-six SPF-grade 8-week-old BALB/c male mice were purchased and randomly divided into the blank group(A Group),the model group(B Group),the loratadine group(C Group),the Wen’s Jian Pi Chu Shi Decoction group(D Group),the Wen’s clearing method group(E Group)and the Wen’s tonifying method group(F Group).The chronic eczema model was constructed by repeatedly stimulating the back of mice with 2,4-dinitrochlorobenzene in acetone solvent.After successful modeling,each group of mice(except for the A Group)was dosed for 14 consecutive days.2.4.2 General situation and skin lesion score before and after modelingRecorded the changes of skin lesions,behavioral activities,mental state,hair,food intake and other general conditions of mice in each group during the process of modeling and drug administration.The severity of skin lesions on the back of mice was quantitatively evaluated after modeling and on the day of experimental sampling.2.4.3 Weight difference and spleen indexMice weight before the experiment and mice weight and spleen weight on the day of sampling were weighed.Calculated the mice body weight difference and spleen index.2.4.4 Histopathological changes of skin lesions on the backHistopathological changes on the dorsal lesions of each group of mice were detected by HE staining.2.4.5 Test of the average optical density values of IL-23,IL-17,FLG and CLDN1 on dorsal lesionsThe expression of IL-23,IL-17,FLG and CLDN1 on the dorsal skin lesions of each group of mice was tested by immunohistochemistry.The AOD value of positive substances was collected and quantified by Image-Pro Plus Image analysis system.2.4.6 Test of IL-23,IL-17,FLG and CLDN1 m RNA on dorsal lesionsThe expressions of IL-23,IL-17,FLG and CLDN1 m RNA on the dorsal lesions of each group of mice were measured by RT-PCR.2.4.7 Test of serum IL-23,IL-17 and Ig EThe serum levels of IL-23,IL-17 and Ig E in each group of mice were measured by ELISA.3.Results3.1 The Characteristics of Chinese herbal medicine in chronic eczema clinical trials in China and its similarities and differences with Sichuan Wen’s school of dermatology The etiology and pathogenesis of chronic eczema was root vacuity and tip repletion.Qi deficiency was the root cause.Wind,moisture and heat were pathological factors.Chronic eczema clinical trials used mainly clear and tonic medicines,which were cold in nature.Common traditional Chinese medicine syndrome types and recommended medication were basically consistent with the guidelines.Wen’s school of dermatology treated chronic eczema with spreading damp-heat pattern mainly from the lung and large intestine meridians,while the spleen vacuity with damp encumbrance type was treated from the lung and spleen meridians,and the blood vacuity and wind-dryness type was treated from the liver and gallbladder meridians.In addition,Wen’s school of dermatology focuses on "clearing heat and detoxication" and "clearing heat and tranquilization".3.2 Effective ingredients,targets and pathways of Wen’s Jian Pi Chu Shi Decoction A total of 142 key active ingredients of Wen’s Jian Pi Chu Shi Decoction were screened,among which quercetin was the most important.51 key action targets,including STAT3,PPARG,PPARA,TP53,MAPK8,etc.were identified.There are up to 269 signaling pathways involved,with 139 intersecting with loratadine,mainly involved in immune or inflammatory aspects,such as the IL-17 signaling pathway,Inflammatory mediator regulation of TRP channels,JAK-STAT signaling pathway,Th1/Th2/Th17 cell differentiation pathway,etc.3.3 Analysis of clinical efficacy and safety of Wen’s Jian Pi Chu Shi Decoction3.3.1 Clinical efficacy and efficacy evaluationIn FAS and PPS analysis,there were statistically significant differences in EASI score,5-D pruritus score,skin barrier index,serum level of IL-23/IL-17,POEM score,DLQI score and TCM syndrome score before and after treatment in the experimental group(P<0.05).In FAS and PPS analysis,except for TCM syndrome score,serum level of IL-23/ IL-17,there were statistically significant differences in EASI score,5-D pruritus score,skin barrier index,POEM score and DLQI score before and after treatment in the control group(P<0.05).In FAS and PPS analysis,there were statistically significant differences in EASI score,skin barrier index,TCM syndrome score and serum level of IL-23/IL-17 between the experimental group and the control group after 4 weeks of treatment(P<0.05),but there was no statistically significant difference in 5-D pruritus score,DLQI score and POEM score(P>0.05).In FAS analysis,the effective rate was 82.22% in the experimental group and 62.22% in the control group,the difference was statistically significant(P<0.05).3.3.2 Safety evaluationIn SS analysis,there was no significant difference in routine blood test,liver and kidney function between the experimental group and the control group before and after treatment(P>0.05).The adverse reaction rate was 8.89% in the experimental group and 13.33% in the control group.There was no significant difference between the two groups(P>0.05).3.4 Study and analysis on the mechanism of Wen’s Jian Pi Chu Shi Decoction3.4.1 General situation before and after modellingExcept for the A Group,dorsal skin of mice in other groups finally showed dark red to rustlike edematous patches after repeated sensitization,which were rough and hypertrophic,and some of them showed chaps and exudates.From the behavioral point of view,the mice gradually transitioned from jumping up and down,restlessness,squeaking,licking,scratching and not wanting to eat to normal activity and free feeding.At the end of the experiment,the skin lesion scores of each administration group were lower than those of the A Group(P<0.05).The skin lesion scores of the C Group,E Group and F Group were higher than those of D Group(P<0.05).3.4.2 Weight difference and spleen indexThere was no significant difference in body weight between the mice in each group(P>0.05).There were significant differences in spleen index among the groups.The B Group and C Group were higher than the A Group(P<0.05).The D Group,E Group and F Group were lower than the B Group(P<0.05).The D Group and F Group were lower than the C Group,(P<0.05).3.4.3 Histopathological changes of skin lesions on the backThe dermis and epidermis of the A Group mice were normal,intact and clear.The mice in the B Group showed dense or lamellar hyperkeratosis in the epidermis or with blood crust,hypertrophic spinous layer,slight spongiosis.The dermis was infiltrated by inflammatory cells,and some capillaries were formed,dilated,and hyperemic.The epidermal hyperkeratosis,spinous layer hypertrophy and inflammatory cell infiltration in dermis were improved in different degrees in the C Group and all Chinese medicine groups.However,most of the mice were still accompanied by capillary neovascularization,dilatation and congestion.3.4.4 Test of the average optical density values of IL-23,IL-17,FLG and CLDN1 in dorsal lesionsIL-23 and IL-17 AOD values on the dorsal lesions of mice in the B Group and each administration group were statistically higher than those in the A Group(P<0.05).IL-23/IL-17 AOD value on the dorsal lesions of mice in each administration group were statistically lower than that in the B Group(P<0.05).Compared with the A Group,the FLG AOD value on the dorsal lesions of mice in the B Group,C Group and E Group decreased significantly(P<0.05).The FLG AOD value on the dorsal lesions of mice in the D Group and F Group were statistically higher than that in the B Group(P<0.05).The CLDN1 AOD value on the dorsal lesions of mice in the B Group and each administration group were lower than that in the A Group(P<0.05).The CLDN1 AOD value was higher in the D Group than in the B Group(P<0.05).3.4.5 Test of IL-23,IL-17,FLG and CLDN1 m RNA on dorsal lesionsIL-23 and IL-17 m RNA on the dorsal lesions of mice in the B Group and each administration group were statistically higher than that in the A Group(P<0.05).IL-23 and IL-17 m RNA on the dorsal lesions of mice in each administration group were statistically lower than that in the B Group(P<0.05).The level of IL-23 and IL-17 m RNA on the dorsal lesions of mice in the D Group were statistically lower than that in the C Group(P<0.05).Except for the D Group,the expression of FLG m RNA on the dorsal lesions of mice in the B Group,C Group,E Group and F Group were lower than that in the A Group(P<0.05).Compared to the B Group,FLG m RNA on the dorsal lesions of mice in the C Group,D Group and F Group were higher(P<0.05).Moreover,the FLG m RNA expression in the D Group was higher than that of the E Group or F Group(P<0.05).The expression of CLDN1 m RNA on the dorsal lesions of mice in the B Group and administration groups were lower than that in the A Group(P<0.05).Except for the A Group,the CLDN1 m RNA on the dorsal lesions of mice in the D Group were higher than that in the other groups(P<0.05).3.4.6 Test of Serum levels of IL-23,IL-17 and Ig ECompared to the A Group,the levels of IL-23 and IL-17 in serum of mice in the B Group,C Group and F Group were significantly higher(P<0.05).The levels of IL-23 and IL-17 in serum of mice in each administration group were significantly lower than the B Group(P<0.05).The serum level of IL-23 and IL-17 in the D Group were lower than that in the C Group(P<0.05).Moreover,the serum level of IL-17 in the E Group was significantly higher than the C Group(P<0.05).The serum Ig E level of mice in the B Group were statistically higher than those in the A Group(P<0.05).The serum Ig E level were statistically lower in the C Group and D Group than that in the B Group(P<0.05).4.Conclusion4.1 Wen’s Jian Pi Chu Shi Decoction could effectively reduce the area and severity of eczema,pruritus,and serum level of IL-23 or IL-17 in chronic eczema patients with spleen deficiency and dampness encumbrance.It also improved patients’ skin barrier,quality of life and TCM symptoms.It was significantly superior to loratadine in terms of EASI score,5-D pruritus score,skin barrier index and TCM symptoms score.4.2 Wen’s Jian Pi Chu Shi Decoction had strong immunosuppressive effect and could reduce IL-23/IL-17 expression in mice serum or dorsal skin lesions.4.3 Wen’s Jian Pi Chu Shi Decoction could exert anti-allergic effect by inhibiting serum Ig E in mice.4.4 Wen’s Jian Pi Chu Shi Decoction repaired the skin barrier,which was manifested by increasing the expression of FLG and CLDN1 on the dorsal skin lesions of mice.4.5 Wen’s Jian Pi Chu Shi Decoction up-regulated the expression of FLG and CLDN1 on dorsal lesions of mice better than its dismantling prescription.The theory of " combination of clearing method and tonifying method" was especially important in repairing the skin barrier.