| Objective: To discuss the TCM theory of the nodular goiter,to improve the level of clinical treatment;Through the study on the clinical application and prescription of the Huo Xue Xiao Ying fang to improve the ability of clinical application;To explore the specific target of its role through the regulation of Huo Xue Xiao Ying fang on the PI3K/Akt signaling pathway.Methods:(1)Theoretical study:The TCM name,etiology,pathogenesis and treatment of nodular goiter were summarized through the review of ancient and modern literature.(2)The characteristics of clinical application research on Huo Xue Xiao Ying fang: we summarized it from the prescription characteristics,the efficacy of each drug and compatibility characteristics,study on pharmacology and technology and clinical application rules.(3)In the experimental part: 60 male and SD rat of SPF grade were randomly divided into 6 groups,fed in the SPF laboratory under the enviroment of constant temperature and humidity,cage rearing.After 2 weeks of feeding,the 0.1% PTU solution was given for a period of 8 weeks.From the beginning of the 9th weekand,the drug was administered continuously for a period of 4 weeks;Lately,the rats were anesthetized by intraperitoneal injection.The blood was collected from carotid artery and the serum was separated,the level of serum TGF-? was detected by Elisa method;The thyroid tissue were isolated: a part of the left thyroid gland was used to make paraffin sections and used to detect apoptosis by TUNEL,part of them were used to detect the expression of PI3 KmRNA,AktmRNA and mTORmRNA by qRT-PCR;Western-Blot was used to detect PTEN,EGFR,ERK1/2,BAX,CyclinD1,FOXO3 a protein expression in the right thyroid gland.The data were collected and analyzed statistically.Results:(1)Theoretical research: The solid thyroid nodules could be named “Yin Jie”,and the nodular goiterwhich was mainly caused by goiter,could be called “Jie Yin”.Qi deficiency was the internal cause of nodular goiter,emotional imbalance was the inducement of nodular goiter,irregular daily life was an important reason for the high incidence of nodular goiter,phlegm and blood stasis was throughout the entire course of nodular goiter,and it was an important pathological factor,a potential cause for the refractory and relapse.The Phlegm Blood Stasis phlegm and blood stasis had the general characteristics and its own characteristics in nodular goiter.The main clinical manifestations were: goiter,thyroid pain,thyroid nodules.In the different diseases,the performance was different;In the secondary or complications,the performance was different;In the varies of severity of the disease,the performance was also different.The Phlegm Blood Stasis of the nodular goiter was mainly in the following forms: liver qi stagnation,phlegm and blood stasis;burn flourishing,phlegm and blood stasis;spleen deficiency,phlegm stagnation and blood stasis;Yin deficiency,phlegm and blood stasis;Yang deficiency,phlegm and blood stasis in the students.At the time of treatment,we should: make clear the severity level of phlegm and blood stasis,regulate emotion and keep yourself happy first,pay attention to diet taboo and cautious on living,pay attention to treatment of the accompanied symptom,treat differently with the compatibility,treat with internal and external combination therapy,treat the chronic disease slowly,prevent the disease recurrence and change.From phlegm and stasis,we summarized eight treatment: Regulating qi-flowing for eliminating phlegm,promoting blood circulation for removing blood stasis;Resolving Phlegm and Moving Blood-stasis,Xiao Ying;Clearing heat and transforming phlegm,activating blood circulation to dissipate blood stasis;Nourishing yin andresolving phlegm and activating blood;Warming yang and resolving phlegm and activating blood;resolving phlegm and activating blood and resolving hard lump;Ventilating the lung and resolving phlegm and resolving hard lump.In the clinical practice,we should analysis carefully for specific diseases,use it flexibly.(2)Huo Xue Xiao Ying fang was applied to nodular goiter with the course was less than 6 months,hypoechoic cystic,clear boundary,no blood supply or low blood supply,the maximum diameter less than 3 cm,a treatment for 3-6 months was appropriate.(3)In the term of histology pathology: The model group showed that: the follicular was swelling and had different size,small part of vesicle fused into bullae,the most was smaller;The arrangement of epithelial cells was irregular,which was developed from the single layer epithelium to the stratified epithelium;The follicular epithelium had a complex structure with nipple like extension;The local follicular hyperplasia was obvious,and the local mass was formed and the surrounding blood vessels were abundant,the copy of the model was successful;After intervention of the drug,the swelling and papillary structure of thyroid follicular in the rats decreased,the surrounding hyperplasia and angiogenesis decreased too,the epithelial cells arrangement were more regular,and with the increase of the concentration of Huo Xue Xiao Ying fang,the improvement was more obvious.(4)The apoptosis index detected by TUNEL method showed that: Compared with the blank control group,the apoptosis index of the model group was significantly lower than that of the control group,and the difference was statistically significant(P<0.01).Compared with the model group,both the western medicine intervention group and all doses of Huo Xue Xiao Ying fang group,the apoptosis index of rat thyroid tissue significantly increased,the differences were statistically significant(P<0.01).Compared the western medicine intervention group with the Chinese medicine group,the difference of apoptosis index between western medicine group and high dose of Huo Xue Xiao Ying fang group was not significant(P>0.05),but it was significantly higher than the low dose and middle dose of Huo Xue Xiao Ying fang groups,the differences were statistically significant(P<0.01).In the each dose group,the apoptosis index changed with the different dose: the high dose group > the middle dose group>the low dose group,the differences were statistically significant(P<0.01).(5)qRT-PCR method was uesd to detect the expression of mRNA in each group of rats: Compared with the blank control group,the expression of PI3 KmRNA,AktmRNA and m TORmRNA in the model group increased,the differences were statistically significant(P<0.01).Compared with the model group,in the middle and high dose of Huo Xue Xiao Ying fang group,the expression of PI3 KmRNA,AktmRNA and mTORm RNA decreased,the differences were statistically significant(P<0.01);There was no significant difference between the western medicine intervention group and the low dose group of Huo Xue Xiao Ying fang(P>0.05).Compared with the Chinese medicine intervention group,the western medicine intervention group had no significant difference with the low dose group of Huo Xue Xiao Ying fang(P>0.05),but there were significant differences between the high and low dose groups(P<0.01).Comparison in the group of TCM: the high dose group was better than the middle dose group,the middle dose group was better than the low dose group,the differences were statistically significant(P<0.01).(6)Protein expression were detected in the thyroid tissue of rats by Western-Blot: Compared with the blank control group,the protein expression of PTEN,BAX and FOXO3 a decreased in the model control group with significant statistical difference(P<0.01).Compared with the model group,in the middle and high dose of Huo Xue Xiao Ying fang group,the expression of PTEN,BAX and FOXO3 a protein increased,the differences were statistically significant(P<0.01);There was no significant difference between the western medicine intervention group and the low dose group of Huo Xue Xiao Ying fang(P>0.05).Compared with the Chinese medicine intervention group,the western medicine intervention group had no significant difference with the low dose group of Huo Xue Xiao Ying fang(P>0.05),there were significant differences between the high and low dose groups(P<0.01).Comparison in the group of TCM: the high dose group was better than the middle dose group,the middle dose group was better than the low dose group,the difference was statistically significant(P<0.01).Compared with the blank control group,the expression of EGFR,ERK1/2 and CyclinD1 protein in the model group increased,the difference were statistically significant(P<0.01).Compared with the model group,in the middle and high dose of Huo Xue Xiao Ying fang group,the expression of EGFR,ERK1/2 and CyclinD1 protein decreased,the differences were statistically significant(P<0.01);There was no significant difference between the western medicine intervention group and the low dose group of Huo Xue Xiao Ying fang(P>0.05).Compared with the TCM intervention group,the western medicine intervention group had no significant difference with the low dose group of Huo Xue Xiao Ying fang(P>0.05),there were significant differences between the high and low dose groups(P<0.01).Comparison in the group of TCM: the high dose group was better than the middle dose group,the middle dose group was better than the low dose group,the differences were statistically significant(P<0.01).(7)Elisa method was used to detect the level of serum TGF-? in rats: compared with the blank control group,the level of serum TGF-? in the model group increased,the difference was statistically significant(P<0.01).Compared with the model group,in the middle and high dose of Huo Xue Xiao Ying fang group,the level of serum TGF-? decreased,the differences were statistically significant(P<0.01);There was no significant difference between the western medicine intervention group and the low dose group of Huo Xue Xiao Ying fang(P>0.05).Compared with the TCM intervention group,the western medicine intervention group had no significant difference with the low dose group of Huo Xue Xiao Ying fang(P>0.05),there were significant differences between the high and low dose groups(P<0.01).Comparison in the group of TCM: the high dose group was better than the middle dose group,the middle dose group was better than the low dose group,the differences were statistically significant(P<0.01).Conclusion:(1)The theoretical system of nodular goiter was perfect,and the pathogenesis theory of phlegm and blood stasis was a great guiding significance for clinical treatment.(2)There was a resistance to apoptosis in the nodular goiter,and the abnormal activation of P13K/Akt signaling pathway may be the specific mechanism of resistance to apoptosis.(3)The principle-method-recipe-medicines of Huo Xue Xiao Ying fang was perfect,the characteristic was obvious,and it was worthy of further popularization and application.(4)Huo Xue Xiao Ying fang can induce apoptosis by regulating the expression of related genes and proteins in the P13K/Akt signaling pathway,and then played a role in the treatment and prevention of malignant transformation:It can prevent malignant nodules by increasing expression of PTEN protein,with the rise of the serum TGF-? levels,the activation of EGFR can inhibit the activation of the PI3K/Akt signaling pathway which regulating the raise of FOXO3 a protein expression,then can inhibit the activity of CyclinD1,the cell growth and proliferation delayed;It also can promote the expression of BAX protein and induce apoptosis;With down regulating expression of ERK1/2 protein,it inhibited the activity of CyclinD1,activated the expression of FOXO3 a and inhibited the proliferation;With down regulation of the expression of mTOR gene,it can inhibit cell proliferation from inhibiting the translation of proteins associated with proliferation. |
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