| Purpose:To observe the effect of eczema No.1 recipe on skin inflammation,barrier function and immune function in acute eczema model of spleen deficiency type guinea pigs,and to explore the mechanism of Chinese medicine?eczema 1 prescription?on acute eczema model of spleen deficiency type guinea pig.To provide clinical basis for the treatment of acute eczema with spleen deficiency.Material and method:1 Materials:Sixty guinea pigs were randomly divided into 6 groups?10 guinea pigs in each group?: blank control group,model group,low dose group of traditional Chinese medicine for acute eczema due to spleen deficiency,high dose group of traditional Chinese medicine for acute eczema due to spleen deficiency,and positive control group of traditional Chinese medicine?Huangbai capsule?.Western medicine?loratadine tablets?positive control group.In the high and low dosage group,traditional methods were used to extract and concentrate,to make decoction,and to be administered by gavage,while in the positive control group,Huangbai capsule was used to dissolve it into suspension with distilled water,and the drug was administered intragastric.The western medicine positive control group was treated with loratadine tablets,dissolved with distilled water to make suspension,and the rest group was gavaged with the same amount of normal saline.2 methods:2.1 methods of modeling: Because TCM syndrome differentiation of eczema is mainly spleen deficiency,this experiment mainly focuses on acute eczema model of guinea pig with spleen deficiency.First,5%DNCB?dinitrochlorobenzene?acetone solution 25 ? L was applied to the cervical hair removal to sensitize the skin.Then the acute eczema model was established by repeated low dose DNCB smear method.On this basis,the spleen deficiency model was established by the method of deficiency of spleen caused by diet and excessive fatigue.Finally,the animal model of acute eczema of guinea pigs with spleen deficiency combined with disease and syndrome was formed.2.2 Experimental methods:Experiment1: effect of eczema No.1 recipe on skin inflammation and barrier function in acute eczema model of guinea pig with spleen deficiency?1?The effect of Formula 1 of eczema on the ear swelling and the difference of ear thickness in the Acute eczema Model of Pigs with spleen deficiency: 1 day after the last stimulation,the guinea pigs were killed,the auricles of both sides of the guinea pigs were cut off,and the two ears were perforated with the metal perforator of diameter 0.9cm in the same part of both ears.The weight of the left and right ear tablets was measured by the electronic balance,and the mass difference was calculated as an indicator of the degree of swelling and the intensity of the inflammatory reaction.?2?The effect of Formula 1 of eczema on the expression of aquaporin-3 in the skin of the acute eczema model of the guinea pig with spleen deficiency: immunohistochemical method was used to detect the expression level of AQP3 in the skin of the guinea pig back,and the computer image analysis system was used to analyze the expression of aquaporin 3.According to the average optical density of positive cells expressed in unit area,the expression of positive cells in tissues was determined.Experiment 2: effect of eczema No.1 recipe on immune function of Acute eczema Model of Pigs with spleen deficiency?1?Effect of eczema No.1 on ECPs in Guinea Pig Serum after 14 days,blood was collected from abdominal aorta and serum was collected after centrifugation before the guinea pig model was killed.Enzyme linked immunosorbent assay?Elisa?was used to determine the content of ECP in the serum of guinea pigs in each group,and the procedure was carried out strictly according to the instructions of the kit.?2?The effect of Formula 1 of eczema on CD4 Tet CD8 T and its ratio in Acute eczema Model of Pig with spleen deficiency: 2 h after the last gastric perfusion,the whole blood of guinea pig was collected by cardiac puncture and anticoagulant,and the specific steps were performed strictly according to the operation process of flow cytometry.The levels of CD4 T 8 T in whole blood of guinea pigs were measured and their ratios were calculated.Results:Experiment 1: effect of eczema No.1 recipe on skin inflammation and barrier function in acute eczema model of guinea pig with spleen deficiency1.Eczema No.1 can reduce the swelling degree of skin and ear,reduce the thickness of ear and reduce the inflammatory reaction in acute eczema guinea pigs with spleen deficiency.Compared with the blank control group,the guinea pigs in the model group had different degrees of ear swelling and the difference of ear thickness was increased.Compared with the model group,all the groups were able to reduce the degree of ear swelling and the difference of ear thickness,the difference was statistically significant?P < 0.01?.The difference between loratadine group and Huangbai capsule group was statistically significant?P < 0.05 or P < 0.01?,and the curative effect was better than that in Huangbai capsule group,but there was no significant difference between the high and low dosage groups of traditional Chinese medicine and that of loratadine group?P > 0.05?.2.Eczema 1 can decrease the overexpression of AQP3 in skin lesions of acute eczema guinea pigs with spleen deficiency.AQP3 protein was expressed in different degree in the skin tissues of each group?light brown to brown,and weakly positive in normal tissues?.Compared with the blank control group,the average optical density in the model group was significantly higher than that in the model group?P < 0.01?,and the average optical density in each treatment group was significantly lower than that in the model group.The difference was statistically significant?P < 0.01?,and there was no significant difference in the mean optical density between the high dose group and loratadine group?P > 0.05?.Compared with the blank group,the expression of AQP3 protein in the model group was strongly positive,the brown granules were distributed in the cell membrane and cytoplasm,and the staining was obviously deepened.Compared with the model group,the expression of AQP3 protein in the treatment group decreased in different levels and the positive products decreased in different degrees.There was a negative correlation between the amount of brown deposition and the dose of eczema 1.Experiment 2: effect of eczema 1 on immune function of acute eczema model in guinea pigs with spleen deficiency1.The formula of eczema 1 can effectively inhibit the overexpression of ECP in serum of acute eczema guinea pigs with spleen deficiency.Compared with the blank control group,the serum ECP in the model group was significantly higher than that in the control group?P < 0.01?,which indicated that the model was successful.Compared with the model group,the serum ECP content in each treatment group was significantly decreased?P < 0.05 or P < 0.01?.The therapeutic effect of high dose Chinese medicine group was better than that of low dose group and Huangbai capsule group?P < 0.05 or P < 0.01?.There was no significant difference between high dose group and loratadine group?P > 0.05?.2.Eczema 1 can balance T lymphocyte subsets by decreasing CD8 T lymphocytes and increasing CD4 T lymphocytes in acute eczema guinea pigs with spleen deficiency.The level of CD4 T lymphocytes in the model group was significantly lower than that in the blank control group?P < 0.01?.The levels of CD4 T lymphocytes in high dose group and loratadine group were all increased?P < 0.05?,but there was no significant difference between high dose group and loratadine group?P > 0.05?.Compared with the model group,the level of CD4 T lymphocytes in the other treatment groups showed an upward trend,but the change was not significant?P > 0.05?.The level of CD8 T lymphocytes in the model group was significantly higher than that in the blank control group?P < 0.01?.The level of CD8 T lymphocytes in high dose group and loratadine group decreased?P < 0.05?,but there was no significant difference between high dose group and loratadine group?P > 0.05?.There was a decreasing trend between the other treatment groups and the model group,but the change was not significant.There was no significant difference?P > 0.05?in CD4 / CD8 ratio between the model group and the normal group?P < 0.01?,and there was an increasing trend between the high dose group and the model group?P < 0.05?,but there was a significant difference between the high dose group and the loratadine group.The difference was not statistically significant?P > 0.05?,but there was no significant difference between the other treatment groups and the model group?P > 0.05?.Conclusion:1.Eczema 1 can reduce the inflammatory reaction and tissue injury of acute eczema guinea pigs with spleen deficiency.2.Eczema 1 can down-regulate the overexpression of AQP3 in the skin lesions of acute eczema guinea pigs with spleen deficiency,so as to alleviate the damage of skin barrier function.3.The formula of eczema 1 can effectively inhibit the overexpression of ECP in serum of acute eczema guinea pigs with spleen deficiency.4.Under the specific dosage and course of treatment,eczema No.1 could decrease the level of CD8 T lymphocytes and increase the level of CD4 lymphocytes in acute eczema guinea pigs with spleen deficiency,thus achieving balance between T lymphocyte subsets. |
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