| Objective: Under the guidance of TCM dialectical theory,through studying the influence of the tutor's experience on the Model rats of hypothyroidism,the effect and mechanism of Yiqi Huatan Huoxue Decoction on the Model rats of hypothyroidism were explored from the angle of TLRs/p38 MAPK signal transduction pathway and intestinal flora.At the same time,Professor Gao Tianxu's experience in clinical treatment of the disease is summarized,providing experimental and theoretical basis for the treatment of hypothyroidism.Materials and methods: 1 Animal experimentsSPF grade 40 2-month-old male Wistar rats were randomly divided into 4 groups: respectively,the normal group(Con),model group(MOD),Positive drug group(LT4),Chinese Medicine Group(ZYH),each group of 10.After 1 week of adaptive feeding,the rat model of hypothyroidism was established by 0.1% Propylthiouracil in 4 weeks.After the success of the mold,according to the daily dose of 60 kg per kilogram of adult rats dosage of dose conversion,the rats were treated by intragastric administration daily regular for 8 weeks.The complete blood count was detected with automated hematology analyzer;the level of serum TT4 was detected with RIA;the level of serum TSH was detected with IRMA,HE staining was used to observe the tissue and morphology of large intestine,immunohistochemical method and Western Blot method was used to observe the proteins related to the signal transduction pathways and the expression of interleukin-1?,16 S r DNA was used to detect the community structure of intestinal flora.2 Experience Summary To summarize the experience of Professor Gao Tianshu's in clinical treatment of hypothyroidism,and to explain the TCM disease name,etiology,pathogenesis,clinical treatment and typical cases,and to provide theoretical basis for the treatment of hypothyroidism with Yiqi Huatan Huoxue Decoction.Results: 1 Based on TLRs/p38 MAPK signal transduction pathway to study the effect of Yiqi Huatan Huoxue Decoction on the large intestine of hypothyroidism in rats 1.1 Rats in generalThe mental state of the Con group rats was good,the reaction was sensitive,the amount of food was normal and the weight was normal.In the Mod group,the rats were slow,unresponsive,with sparse fur,decreased consumption and stool.The LT4 group and the ZYH group is consistent with the Mod group after the modeling and the situation improved after treatment,which was close to the Con group.1.2 Routine results of peripheral blood bloodrats showed that leukocyte count(WBC),neutrophil proportion(%NEUT),percentage of Eosinophil(%EOS),percentage of neutrophils(%BASO),percentage of monocytes(%MONO),percentage of lymphocytes(%LYMPH),platelet count(PLT)in each group of rats None of the results were statistically significant(p>0.05).Red blood cell count(RBC),hemoglobin(HB)in Mod,LT4,ZYH group were lower than the Con group,the difference was statistically significant(p<0.05).1.3 TSH and TT4 level in rats serumAfter modeling,compared with Con group,the levels of TSH in Mod,LT4 and ZYH three groups were increased,the TT4 level were decreased,and the difference was statistically significant(P<0.05).There was no statistical significance in the three groups(P>0.05),indicating that the model was successful.After treatment,compared with Mod group,the serum TSH level of LT4 and ZYH group decreased and the TT4 level increased,the difference was statistically significant(P<0.05),and the effect of ZYH is better than LT4(P<0.05).1.4 HE staining of large intestine tissue in ratsMicroscopic observation found,the Con group had complete structure and no significant abnormality;There were more inflammatory cells in Mod group,swelling of large intestine gland,bleeding in some intestinal mucosa,and partial shedding of submucosa;The congestion of LT4 and Zyh group decreased obviously and the range of hemorrhage was reduced.1.5 Expression of protein in large intestine tissue of rats 1.5.1 Immunohistochemical detectionThe results showed that TLR4,My D88 and p38 MAPK were all expressed in 4 groups of large intestine,compared with Con group,the light density of mod group increased significantly,and the difference was statistically significant(p<0.05).Compared to the Mod group,The light density value of LT4 and ZYH group decreased obviously,the difference was statistically significant(p<0.05).Il-1? was expressed in 4 experimental groups,compared with con group,the density value of the Mod group was significantly higher,the difference was statistically significant(p<0.05);Compared with mod group,the light density value of LT4 and ZYH group decreased significantly,the difference was statistically significant(p<0.05).1.5.2 Western Blot detectionThe results showed that Compared with Con group,the protein expression of TLR4,My D88 and p38 MAPK in Mod group was enhanced statistically(p<0.05).Compared with Mod group,the expression of protein in LT4 and ZYH group decreased significantly,there was statistically significant(p<0.05),and ZYH group protein expression was lower than that in LT4 group.,statistically significant(p<0.05).Compared with Con group,the expression of IL-1? protein in Mod group was enhanced statistically(p<0.05)and compared with Mod group,the expression of protein in LT4 and ZYH group decreased significantly,there was statistically significant(p<0.05),and ZYH group protein expression was lower than that in LT4 group,which was statistically significant(p<0.05).2 Effect of Jianpi Huatan Huoxue decoction on improving intestinal flora regulating hypothyroidism 2.1 Total abundance of speciesAt the phylum level,the highest degree of abundance were Firmicutes,Bacteroidetes,Euryarchaeota and proteobacteria,which occupies more than 90%.At the genus level,the highest abundance were lactobacillus,methanosphaera,Bacteroides and Faecalibaculum.2.2 Multi-sample species compositionAt the phylum level,compared with the Con group,the proportion of Euryarchaeota in the Mod group was increased and the proportion of Bacteroidetes was reduced(P<0.05).Compared with the Mod group,the proportion of Firmicutes in LT4 and ZYH groups decreased and the proportion pf Bacteroidetes and Proteobacteria were increased(P<0.05),and there was no significant difference between the two groups of LT4 Group and ZYH Group(P>0.05).2.3 Diversity differences of flora Compared with the con group,the diversity of the Flora in Mod group decreased and the diversity of LT4 and ZYH groups increased,and the diversity of the two groups was higher than that in the Con group.2.4 Correlation analysis of species and environmental factorsAt the genus level,the level of TSH was positive correlated with Anaerobiospirillum,Eubacterium,Akkermansia,Methanosphaera,Kitasatospora and Rothia;were negative correlated with turicibacter,facklamia,Corynebacterium,Jeotgalicoccus and Desulfovibrio.The level of TT4 was positively correlated with Turicibacter,facklamia,Corynebacterium,Jeotgalicoccus and Psychrobacter.Conclusion: 1 Yiqi Huatan Huoxue Decoction can improve the thyroid function of the model rats with hypothyroidism,regulate TSH,TT4 level and reduce the inflammation state of the large intestine,suggesting that the decoction may have the function of improving the thyroid function by alleviating the inflammation state,and the effect is better than LT4.2 Yiqi Huatan Huoxue Decoction can regulate the TLRs/p38 MAPK signal transduction pathway in the rat model of hypothyroidism,reduce the expression of TLR4,My D88 and p38 MAPK protein,and decrease the secretion of IL-1?.3 Yiqi Huatan Huoxue Decoction can effectively improve the intestinal flora composition of the model rats with hypothyroidism,increase the species diversity,regulate the intestinal flora and then regulate the thyroid function,which may be one of the mechanisms to improve the hypothyroidism.4 Professor Gao is based on the different stages of thyroid dysfunction,put forward the "invigorating Qi and invigorating spleen method" and "invigorating spleen and reducing phlegm and invigorating the circulation of blood" of treatment ideas,to obtain a better clinical effect. |
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