| Objective:To observe the mechanism of estrogen receptor and PI3K/Akt/mTOR pathway affecting the precancerous lesions of breast cancer.The molecular mechanism of Shugan Jianpi Jiedu decoction can reverse the progression of breast cancer in rats by regulating ER’s influence on PI3K/Akt/mTOR pathway expression.Methods:SD rats were taken and divided into 7 groups(10 rats in each group),namely blank group,breast precancerous lesion model group,shugan jianpi jiedu decoction with low dose group,shugan jianpi jiedu decoction with middle dose group,shugan jianpi jiedu decoction with high dose group,tamoxifen group-western medicine control group and Xihuang capsule group-traditional Chinese medicine(TCM)control group.DMBA modeling method was used to carry out modeling forbreast precancerous lesion model group,shugan jianpi jiedu decoction with low dose group,shugan jianpi jiedu decoction with middle dose group,shugan jianpi jiedu decoction with high dose group,tamoxifen group-western medicine control group,Xihuang capsule group-TCM control group.In shugan jianpi jiedu decoction with low dose group,shugan jianpi jiedu decoction with middle dose group and shugan jianpi jiedu decoction with high dose group,the above decoction was given by intragastric administration on the 2nd day after modeling.In tamoxifen group-western medicine control group,tamoxifen was given by intragastric administration on the 2nd day after modeling.In Xihuang capsule group-TCM control group,Xihuang capsule was given by intragastric administration on the 2nd day after modeling.In blank group,sesame oil was given by intragastric administration,withnatural feeding.After 4 weeks,anesthesia was performed and materials were taken.The body weight and related organ coefficients were calculated,and the pathological changes of mammary gland and ovarian tissues were detected by HE staining.The protein expression of ER,PI3K,Akt and mTOR was detected by immunohistochemistry and Western blot,and the mRNA expression of ER,PI3K,Akt and mTOR was detected by RT-PCR;IL-2,IL-4,IL-6,IL-10,IL-12,FSH,LH,PRL,P,T and E2 were detected by ELISA;CD3+,CD4+,CD8+and CD4+/CD8+were detected by flow cytometry.IgA,IgG and IgM were detected by immuno-turbidimetric method.Results:1.Pathological changes:In the model group,breast tissue showed a mass of hyperplasia of breast acinus and ducts,with different sizes and shapes,dense acinus in lobules,obvious expansion of glandular duct cavity and acinar cavity,and a mass of secretions in the cavity.Proliferative cells filled the duct cavity,and the phenomenon of overlapping and pushing appeared between cells,and the number of mammary epithelial cells increased;immunohistochemistry,Western blot and RT-PCR showed that the expression of ER,PI3K,Akt and mTOR in the model group was higher than those in the blank group.2.The body weight inbreast precancerous lesion model group decreased,and gradually increased from shugan jianpi jiedu decoction with low dose group to Xihuang capsule group-TCM controlgroup;compared with the blank group,the coefficients of ovary and uterus in precancerous lesion model group,shugan jianpi jiedu decoction with low dose group,shugan jianpi jiedu decoction with middle dose group,shugan jianpi jiedu decoction with high dose group and Xihuang capsule group-TCM control group increased significantly,and the difference was statistically significant(P<0.05);compared with the breast precancerous lesion model group,the coefficients in shugan jianpi jiedu decoction with low-dose group,shugan jianpi jiedu decoction with middle-dose group,shugan jianpi jiedu decoction with high-dose group,tamoxifen group-western medicine control group and Xihuang capsule group-TCM control group decreased significantly,and the difference was statistically significant(P<0.05).3.The expression of ER,PI3K,Akt and mTOR detected by immunohistochemistry,western blot and RT-PCR showed that shugan jianpi jiedu decoction had inhibitory effect on the expression of ER,PI3K,Akt and mTOR.Among them,shugan jianpi jiedu decoction with high-dose group had the best effect,followed by tamoxifen group-western medicine control group.Correlation analysis was used to study the correlation between ER and PI3K,mTOR as well as Akt.PI3K(P=0.667)>0.05 was obtained,showing that there is no obvious correlation between ER and PI3K.The correlation coefficient between ER and mTOR as well as Akt was 1.000,and there was a significant difference between ER and Akt(P<0.01),which shows that there is a significant positive correlation between ER and Akt.4.In terms of CD3+,CD4+,CD8+ and CD4+/CD8+detected by flow cytometry,CD3+ in shugan jianpi jiedu decoction with low dose group was significantly higher than that in other groups(P<0.05);the expression of CD4+ and CD8+in breast precancerous lesion model group was lower than that in other groups(P<0.05),the expression of CD4+and CD8+was the highest in shugan jianpi jiedu decoction with low dose group;IgA,IgG and IgM in breast precancerous lesion model group were lower than those in other groups(P<0.05),but higher in shugan jianpi jiedu decoction withmiddle dose group,shugan jianpi jiedu decoction with high dose group,tamoxifen group-Western medicine control group and Xihuang capsule group-TCM control group(P<0.05);ELISA showed that IL-2,IL-4,IL-6,IL-10 and IL-12 levels in the model group were lower than those in the other groups(P<0.05),and the highest levels in the Xihuang capsule-TCM control group.The levels of IL-6 and IL-12 inbreast precancerous lesion model group were higher than those in other groups(P<0.05),and the lowest in the shugan jianpi jiedu decoction with high dose group.5.The expression of FSH,LH,PRL,P,T and E2 inbreast precancerous lesion model group was higher than that in other groups(P<0.05),and the expression of FSH,LH,PRL,P,T and E2 in LSDP with high dose group was the lowest,which was similar to that in tamoxifen group-western medicine control group.Conclusion:1.ER is positively correlated with Akt/mTOR signaling pathway,and the changes of gonadal axis are found in breast precancerous lesion,which may be caused by the increase of ER through the action of gonadal axis,thus causing the increase of PI3K/Akt/mTOR expression.2.Shugan jianpi jiedu decoction may inhibit the expression of ER by inhibiting the gonadal axis,thus inhibiting the expression of PI3K/Akt/mTOR signaling pathway.Meanwhile,it can reverse the breast precancerous lesion by regulating immunity and enhancing immune response. |
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